The Molecular Pathology of Schizophrenia: An Overview of Existing Knowledge and New Directions For Future Research

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The molecular pathology of schizophrenia: an overview of

existing knowledge and new directions for future research
1 1,2 ✉
Takumi Nakamura and Atsushi Takata
© The Author(s) 2023
Despite enormous efforts employing various approaches, the molecular pathology in the schizophrenia brain remains elusive. On
the other hand, the knowledge of the association between the disease risk and changes in the DNA sequences, in other words, our
understanding of the genetic pathology of schizophrenia, has dramatically improved over the past two decades. As the
consequence, now we can explain more than 20% of the liability to schizophrenia by considering all analyzable common genetic
variants including those with weak or no statistically significant association. Also, a large-scale exome sequencing study identified
single genes whose rare mutations substantially increase the risk for schizophrenia, of which six genes (SETD1A, CUL1, XPO7, GRIA3,
GRIN2A, and RB1CC1) showed odds ratios larger than ten. Based on these findings together with the preceding discovery of copy
number variants (CNVs) with similarly large effect sizes, multiple disease models with high etiological validity have been generated
and analyzed. Studies of the brains of these models, as well as transcriptomic and epigenomic analyses of patient postmortem
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tissues, have provided new insights into the molecular pathology of schizophrenia. In this review, we overview the current
knowledge acquired from these studies, their limitations, and directions for future research that may redefine schizophrenia based
on biological alterations in the responsible organ rather than operationalized criteria.
Molecular Psychiatry; https://doi.org/10.1038/s41380-023-02005-2
INTRODUCTION the critical problem of multiple testing and relevant statistical

The term pathology is defined as the study of the essential nature power. This has especially been true in comprehensive studies of
of disease [1]. In the pathology of physical diseases, abnormal variants in DNA, which encodes fundamental biological informa-
changes in responsible organs or systems, such as invasion of tion for all organs, including the brain and can be analyzed by
malignant cells in cancers, and loss of nigrostriatal dopaminergic using easily accessible peripheral tissue samples, resulting in an
neurons in Parkinson’s disease, are examined typically by accumulation of statistically robust findings. Given these, while we
microscopes. On the other hand, a field of study of pathological should refrain from being overly optimistic, perhaps now might be
cognition and behaviors often observed in patients with the time to bring together these technologies and resources to
psychiatric disorders, psychopathology, does not analyze the elucidate the molecular pathology of schizophrenia. In this review,
brain, the organ presumably responsible for these disorders. This we overview the findings from research aiming to decipher the
may not be unreasonable given that psychiatry is etymologically a molecular pathology of schizophrenia, emphasizing large-scale
study of the psyche, a Greek word ψυχή whose derived meaning omics studies with substantial statistical power and analyses of
includes invisible spirit or soul. Nevertheless, It has long been etiologically valid disease models, and summarize the current
thought that mental illness is fundamentally a disease of the brain, achievements. Following that, the challenges and obstacles to be
as classically advocated by Wilhelm Griesinger, Emil Kraepelin, and overcome and the future research directions will be discussed,
others [2, 3]. Based on this concept, the brain pathology of along with an introduction of preliminary results from studies
schizophrenia, a common psychiatric disorder with a lifetime utilizing cutting-edge technologies that will surely facilitate the
prevalence of ~1% [4], has been investigated in numerous studies. elucidation of the fundamental pathology of schizophrenia.
However, despite many efforts, no definite pathological changes,
like senile plaques and neurofibrillary tangles in Alzheimer’s
disease, were identified in their postmortem brains. Meanwhile, THE GENETIC PATHOLOGY OF SCHIZOPHRENIA
recent rapid advances in molecular biology and engineering have Almost unquestionably, the brain is the organ primarily respon-
prompted the development of methods to analyze molecules sible for the pathogenesis of schizophrenia. However, the human
such as nucleotides and proteins more comprehensively, more brain is covered by thick cranial bones, and therefore it is almost
sensitively, with a higher cellular and spatial resolution, and impossible to access and analyze living human brain tissues at the
quantitatively. Besides, multiple collaborative research frameworks molecular level in a non-invasive manner, nor is it easy to collect
have been established to ensure sample sizes sufficient to solve postmortem brains on a large scale. On the other hand, the
1
Laboratory for Molecular Pathology of Psychiatric Disorders, RIKEN Center for Brain Science, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan. 2Research Institute for Diseases of Old
Age, Juntendo University Graduate School of Medicine, 2-1-1 Hongo, Bunkyo-Ku, Tokyo 113-8421, Japan. ✉email: [email protected]
Received: 1 August 2022 Revised: 15 February 2023 Accepted: 15 February 2023

T. Nakamura and A. Takata
2
sequences of DNA, the molecule encoding fundamental biological nucleotide variants (SNVs, any frequency of single nucleotide
information of all tissues, can be analyzed without accessing the substitutions including both SNPs and rare variants) and short
brain, since its sequences are in principle identical in every cell insertion/deletions (indels) in protein-coding regions were system-
and invariant throughout life, with few exceptions. Based on this atically analyzed by using exome (all protein-coding exonic
relative ease in obtaining samples as well as the high heritability regions) sequencing data of 24,248 schizophrenia cases, 97,322
of schizophrenia, reported to be 60–80% in epidemiological controls, and 3402 trios consisting of schizophrenia probands and
studies [5–8], large-scale genetic studies, analyzing samples from their unaffected parents. By evaluating the burden of rare loss-of-
more than 100,000 individuals these days, have been conducted. function (LOF: nonsense, splice site, and frameshift indel) variants
Reflecting this large number, among various research on the (also known as protein-truncating variants: PTVs) and damaging
molecular pathology of schizophrenia, statistically robust findings missense variants (defined by the missense badness, PolyPhen-2,
have been particularly accumulated from human genetics studies and constraint [MPC] score), including those arisen de novo in the
analyzing variations of the sequences of DNA molecules. For this probands, this study identified ten genes (SETD1A, CUL1, XPO7,
reason, we begin with an overview of our knowledge of the TRIO, CACNA1G, SP4, GRIA3, GRIN2A, HERC1, and RB1CC1; Table 1b)
genetic pathology of schizophrenia. surpassing the exome-wide significance threshold (defined as
2.14 × 10−6 based on the number of protein-coding genes
Robust findings from large-scale analyses of common and rare analyzed) and 22 additional genes at a false discovery rate
variants (FDR) < 0.05. Considering their effect sizes, six (SETD1A, CUL1,
Although schizophrenia is a highly heritable disorder, no single XPO7, GRIA3, GRIN2A, and RB1CC1) out of the ten exome-wide
variant explaining a large portion of the overall heritability has significant genes were enriched for rare PTV and damaging
been identified. Therefore, comprehensive studies of various allele missense (MPC > 3) variants with OR > 10, indicating that rare
frequencies and effect sizes of genetic variants contributing to its deleterious variants of these genes confer the schizophrenia risk
risk are warranted. In a simplified view, there are two major types with large effects as well as robust statistical significance. Another
of genetic studies on different frequencies and effect sizes of notable thing is that five (SETD1A, TRIO, CACNA1G, GRIA3, and
variants, that is, genome-wide association studies (GWAS) of GRIN2A) of the ten genes are also implicated in other neurode-
common single nucleotide polymorphisms (SNPs) with small velopmental disorders, such as intellectual disability and epilepsy,
effect sizes (typically odds ratio [OR] < 1.2) and sequencing studies as registered on the Online Mendelian Inheritance in Man
of rare variants potentially with large effect sizes (sometimes (OMIM) database [24]. Therefore, on the one hand, it is possible
OR > 10). The scales of these two types of studies have that these cases with deleterious variants in known neurodeve-
consistently grown [9–19], and the results of the two largest lopmental disorder genes might be individuals who should have
studies to date, each looking into common SNPs [20] or rare been molecularly diagnosed as patients of highly heritable
variants [21], have recently been published after peer review. neurodevelopmental diseases, while on the other hand, another
In the newest GWAS of common SNPs by the Psychiatric possibility is that the carriers of these variants did not exhibit
Genomics Consortium (PGC) analyzing 76,755 schizophrenia cases developmental and physical symptoms sufficient to be diagnosed
and 243,649 control individuals [20], 287 distinct loci with with a neurodevelopmental disease due to some modifying
genome-wide significant association (P < 5 × 10−8) were identified. factors and were operationally diagnosed with schizophrenia. If
The SNP with the largest effect size was the rs140365013 variant the former is true, this suggests that genetic testing may detect
on chromosome 6 near the major histocompatibility complex overlooked patients with highly heritable neurodevelopmental
region, with an OR of 1.23, confirming that individual SNPs do not diseases and provide clues for better intervention. If the latter is
greatly increase the disease risk. On the other hand, the the case, it implies that some modifying factors influence the
proportion of the variance in schizophrenia liability explained by severity of the symptoms. Indeed, there is accumulating evidence
all measured SNPs, including numerous variants that did not show supporting the existence of modifying factors, such as common
statistically significant association with schizophrenia, was and rare variants other than the diagnostic mutation, in patients
reported to be 24%. This proportion is much larger than that with neurodevelopmental diseases [25–28]. A more detailed
calculated solely from loci associated with genome-wide sig- analysis of such modifying factors may pave the way toward the
nificance. Therefore, a significant part of the heritability is development of new treatment and prevention strategies.
attributable to common SNPs that individually show weak Besides common SNPs analyzed in GWAS and rare SNVs/
associations and effects, and it can be said that schizophrenia is indels analyzed in exome sequencing studies, another important
a highly polygenic disorder. Since each of the 287 associated loci class of genetic variation of which several are known to be
often contains multiple genes (specifically, of the 287 loci, 206 and robustly associated with schizophrenia is copy number variants
108 contain ≥2 and ≥5 genes, respectively), it is essential to (CNVs) [29–32]. Among important works on CNVs, a large
identify functionally important causal genes and variants in order genome-wide study by the CNV Working Groups of PGC
to understand the molecular pathology from the genetic analyzing 21,094 schizophrenia cases and 20,227 controls [31]
pathology. To this end, gene prioritization was performed in this identified copy number losses at six loci (1q21.1, 2p16.3
GWAS by PGC using various approaches such as fine-mapping of involving NRXN1, 3q29, 15q13.3, distal 16p11.2, and 22q11.2)
credible sets of causal SNPs by an adaptation of a Bayesian and gains at two loci (7q11.23 and proximal 16p11.2) that are
inference algorithm [22] and Mendelian randomization to identify significantly (P < 1.33 × 10−4 for losses and 4.33 × 10−5 for gains)
SNPs whose causal effects are likely to be mediated through associated with schizophrenia after multiple testing corrections
regulation of gene expression [23]. As a result, a total of 120 (Table 1c). All of these are rare in controls and contribute to the
prioritized genes were nominated, of which 70 and 55 received risk for schizophrenia with ORs ranging from 3.8 to infinity. Also,
support from the fine-mapping and Mendelian randomization like the known neurodevelopmental disorder genes identified in
analyses, respectively. There were five genes supported by both the SCHEMA study described above, all, or nearly all of these
lines of evidence (CUL9, FURIN, LINC00320, SNAP91, and ZNF823). In schizophrenia-associated CNVs are phenotypically pleiotropic
addition, two other prioritized genes (GRIN2A, and SP4) are and often more strongly associated with disorders other than
supported by statistical evidence significant after conservative schizophrenia, such as ASD and intellectual disability. Therefore,
multiple testing correction in the rare variant study described complex phenotype-genotype relationships should be consid-
below (Table 1a, b). ered when predicting disease risks from the information on
In a companion study of rare variants by the Schizophrenia CNVs and generating and analyzing CNV-based animal and
Exome Meta-Analysis (SCHEMA) Consortium, rare single cellular models.
Molecular Psychiatry
Table 1. Prioritized schizophrenia genes from common and rare variant studies.
a. PGC GWAS of Common SNPs (76,755 cases and 243,649 controls, ref. [20])*
Gene Symbol Ensembl gene ID Chromosome Gene biotype Index SNP Position (hg19) Ref Alt Uncorrected Control PGC Prioritized Prioritized by SCHEMA SCHEMA
ID P value frequency odds ratio by fine Mendelian uncorrected odds ratio**
(95% CI) mapping randomization P value (95% CI)
CUL9 ENSG00000112659 6 protein_coding rs113113059 43160375 T C 2.29E-11 0.756 1.06 (1.04-1.08) Yes Yes 0.548 1.03 (0.436–2.19)
FURIN ENSG00000140564 15 protein_coding rs4702 91426560 G A 2.15E-23 0.446 1.08 (1.07-1.10) Yes Yes 0.066 2.68 (0.555–11.3)
GRIN2A ENSG00000183454 16 protein_coding rs9926049 9939960 C A 3.16E-10 0.736 0.95 (0.93-0.96) Yes No 7.37E-07† 24.1 (5.36-221)
LINC00320 ENSG00000224924 21 lincRNA rs459391 22120508 T C 1.54E-08 0.185 1.06 (1.04-1.08) Yes Yes NA NA
SNAP91 ENSG00000065609 6 protein_coding rs2022265 84293271 A G 3.74E-10 0.523 1.05 (1.03-1.07) Yes Yes 1 0 (0–6.08)
SP4 ENSG00000105866 7 protein_coding rs7811417 21534152 T C 2.17E-09 0.347 1.05 (1.03-1.07) Yes No 5.08E-07† 7.59 (3.2-19.3)
ZNF823 ENSG00000197933 19 protein_coding rs72986630 11849736 C T 3.07E-10 0.939 0.89 (0.86-0.93) Yes Yes 0.438 1.38 (0.602–2.92)
b. SCHEMA study of rare SNVs/indels (24,248 cases, 97,322 controls, and 3,402 case trios, ref. [21])
Gene Symbol Ensembl gene ID Chromosome Case LOF/ Control Case missense Control De novo Uncorrected P Bonferroni Odds ratio* OMIM OMIM phenotype
PTV count LOF/ (MPC ≥ 3) count missense LOF/ (95% CI) gene ID
PTV count (MPC ≥ 3) count PTV count
SETD1A ENSG00000099381 16 15 3 3 4 3 2.00E-12 4.66E-08 10.3 (4.12-29.3) 611052 Neurodevelopmental disorder
with speech impairment and
dysmorphic facies
CUL1 ENSG00000055130 7 8 1 2 0 3 2.01E-09 4.69E-05 44.2 (6.42-253) 603134 None
XPO7 ENSG00000130227 8 12 1 1 1 1 7.18E-09 0.000167 28.1 (6.46-253) 606140 None
TRIO ENSG00000038382 5 18 16 0 0 2 6.35E-08 0.001481 5.02 (2.47-10.4) 601893 Intellectual developmental
disorder, autosomal dominant,
with microcephaly
CACNA1G ENSG00000006283 17 10 13 8 4 0 4.57E-07 0.010658 4.25 (2.07-8.78) 604065 Spinocerebellar ataxia, early-
onset, severe, with
neurodevelopmental deficits
SP4 ENSG00000105866 7 13 6 3 3 1 5.08E-07 0.011847 7.59 (3.2-19.3) 600540 None
GRIA3 ENSG00000125675 X 5 0 3 2 1 5.98E-07 0.013946 20.1 (4.28-188) 305915 Intellectual developmental
disorder, X-linked, syndromic,
Wu type
GRIN2A ENSG00000183454 16 9 2 3 0 0 7.37E-07 0.017188 24.1 (5.36-221) 138253 Epilepsy, focal, with speech
disorder and with or without
impaired intellectual
development
HERC1 ENSG00000103657 15 28 32 0 0 0 1.26E-06 0.029384 3.51 (2.04-6.03) 605109 None
RB1CC1 ENSG00000023287 8 9 4 0 0 2 0.000002 0.046642 10 (2.89-43.9) 606837 None
c. PGC CNV Working Group study (21,094 cases and 20,227 controls, ref. [31])
Locus (gene) Chromosome Start (hg18) End (hg18) CNV type Case count Control count Regional Odds ratio Contained genes
uncorrected P value (95% CI)
22q11.21 22 17400000 19750000 Loss 64 1 5.70E-18 67.7 DGCR2, AC004471.2, TSSK2, DGCR14, GSC2, SLC25A1, CLTCL1, HIRA, MRPL40,
(9.3-492.8) C22orf39, UFD1L, CDC45L, CLDN5, SEPT5, GP1BB, TBX1, GNB1L, C22orf29,
TXNRD2, COMT, ARVCF, C22orf25, DGCR8, HTF9C, RANBP1, ZDHHC8, RTN4R,
DGCR6L, AC007663.29, AC023490.5-2, GGTLC3, TMEM191C, PI4KAP1, RIMBP3,
AC011718.2, AC007731.16-3, AC007731.16-4, USP41, ZNF74, SCARF2, KLHL22,
MED15, POM121L4P, PI4KA, SERPIND1, SNAP29, CRKL, AC002470.17-2, AIFM3,
LZTR1, THAP7, AC002472.8-1, P2RX6, SLC7A4, P2RX6P, AC002472.8-2
16p11.2, 16 29560000 30110000 Gain 70 7 2.52E-12 9.4 SPN, QPRT, C16orf54, AC009133.7-1, KIF22, MAZ, PRRT2, C16orf53, MVP, CDIPT,
proximal (4.2-20.9) AC120114.2-2, SEZ6L2, ASPHD1, KCTD13, TMEM219, TAOK2, HIRIP3, INO80E,
DOC2A, AC093512.2, FAM57B, ALDOA, PPP4C, TBX6, YPEL3, GDPD3, MAPK3,
CORO1A, BOLA2B, ZNF688, ZNF688
2p16.3 2 50000992 51113178 Loss 35 3 4.92E-09 14.4 NRXN1
(NRXN1) (4.2-46.9)
15q13.3 15 28920000 30270000 Loss 28 2 2.13E-07 15.6 MTMR15, MTMR10, TRPM1, KLF13, OTUD7A, CHRNA7
(3.7-66.5)
3
4
Aggregating the findings from studies of common SNPs and
PDZK1P2, PDZK1P2, NBPF8, NBPF8, FAM108A3, AL049742.8-2, NBPF12, PRKAB2,
TFRC, ZDHHC19, AC069257.28-2, PCYT1A, TM4SF19, TCTEX1D2, UBXN7, C3orf43,
TRIM50, FKBP6, FZD9, BAZ1B, BCL7B, TBL2, MLXIPL, VPS37D, DNAJC30, WBSCR22,
STX1A, ABHD11, CLDN3, AC093168.3, CLDN4, WBSCR27, WBSCR28, ELN, LIMK1,
RNF168, FBXO45, WDR53, C3orf34, LRRC33, PIGX, PAK2, SENP5, NCBP2, PIGZ,
FMO5, CHD1L, BCL9, ACP6, GJA5, GJA8, GPR89B, PDZK1P2, NBPF11, NBPF11,
CI: confidence interval; LOF/PTV: loss-of-function/protein-truncating variant; OMIM: Online Mendelian Inheritance in Man; MPC: missense badness, PolyPhen-2, and constraint; SNV: single nucleotide variant.
rare SNVs/indels and CNVs, we can now explain a substantial
part of the schizophrenia heritability (mainly by common SNPs)
ATXN2L, TUFM, SH2B1, ATP2A1, RABEP2, CD19, NFATC2IP, SPNS1, LAT,

and have produced a list consisting of six genes and six CNVs
associated with schizophrenia with observed ORs larger than
CI: confidence interval; FDR: false discovery rate; LOF/PTV: loss-of-function/protein-truncating variant; MPC: missense badness, PolyPhen-2, and constraint; SNP: single nucleotide polymorphism.
ten. Also, there is a convergence of the results of studies of
common and rare variants. At the level of individual genes,
GRIN2A and SP4 are included in the list of 120 genes prioritized
in the PGC GWAS and showed exome-wide significant enrich-
WBSCR1, LAT2, RFC2, CYLN2, GTF2IRD1, GTF2I

ment of rare deleterious variants in the SCHEMA study, as
described above. At the level of overall enrichment patterns,
the sets of genes implicated in the SCHEMA study and studies
FAM108A2, FAM108A2, BX842679.19
of rare coding variants in other neurodevelopmental disorders

(e.g., ASD and intellectual disability) are shown to be
significantly enriched for common variant associations in the
PGC GWAS.
We provide a compiled list of genes and CNVs identified
Contained genes
MFI2, DLG1, BDH1
through large-scale studies, which underlie the genetic pathology

AC112166.3-2
of schizophrenia, in Table 1, together with the landscape of their

population frequencies and effect sizes in Fig. 1. Meanwhile, as
shown in Table 1 and Fig. 1, it should be noted that there are wide
ranges of confidence intervals for ORs for rare risk variants. Also,
there is a possibility of the so-called winner’s curse [33] in genetic
Odds ratio
(2.6-162.2)
(3.1-125.7)
(95% CI)
studies. To more accurately estimate their effect sizes and the

(2.1-6.9)
Infinity
robustness of the association, further larger studies are always

20.6
16.1
3.8
warranted.
Functional convergence of the findings in genetic studies

uncorrected P value
As described above, genetic studies have identified a number of

*Results from the "core" meta-analysis downloaded from https://www.med.unc.edu/pgc/download-results/ are shown.
statistically robust new genes and variants; however, these in

themselves only show “association” with the phenotype. Thus,
Regional
0.000168
1.50E-06
1.86E-06
5.52E-05
further analysis is needed to translate genetic findings into the

knowledge of the molecular brain pathology of schizophrenia.
This can be facilitated, for example, by testing if specific
molecular and biological pathways are enriched among the
Control count
associated genes.
More specifically, gene ontology (GO) enrichment analyses
were performed in both the PGC GWAS and the SCHEMA study
14
by utilizing MAGMA [34] and DNENRICH [16] that appropriately

control for confounding factors such as gene sizes and linkage
Case count
disequilibrium. We, therefore, examined how the results of these

two studies converge at the levels of molecular function. In the
PGC GWAS and the SCHEMA study, the results of GO enrichment
60
16
11
16
analysis for 7315 and 1491 terms are available, respectively. Of

these, 1431 GO terms were commonly analyzed and 111 of them
CNV type
c. PGC CNV Working Group study (21,094 cases and 20,227 controls, ref. [31])
+ gain
showed significant enrichment at uncorrected P < 0.05 in both of

Gain
Loss
Loss
Loss
these two studies (Fig. 2a). The statistical significance (−log10 P

value) for each term in the PGC GWAS and the SCHEMA study
showed a highly significant correlation (Fig. 2b, Pearson’s
End (hg18)
**Odds ratio for LOF/PTV and MPC ≥ 3 missense variants.

146176000
198840000
correlation coefficient = 0.39, P = 6.67 × 10−54). This result indi-

*Odds ratio for LOF/PTV and MPC ≥ 3 missense variants.
28960000
73780000
cates that there is a convergence of molecular and biological

CI: confidence interval; CNV: copy number variant.
pathways implicated by common SNPs and rare deleterious

variants. Specifically, four GO terms, all related to voltage-gated
Start (hg18)
channels and synaptic transmissions, were significant after

144646000
197230000
28730000
72380000
Bonferroni correction in both the PGC GWAS and the SCHEMA

study (Fig. 2b). When the 32 GO terms with Bonferroni-corrected
P < 0.05 in either dataset (respectively 25 and 11 terms in the
PGC GWAS and the SCHEMA study, of which four are common as
Bonferroni-corrected P < 0.05.
Chromosome
above) were visualized as networks by connecting them based

on the similarity of the contained genes (Fig. 2c), we observed
the formation of three clusters, each related to channel or
continued
16
transporter activities; neuronal components (synapse, axon, and

1
dendrite); chromatin or histone organization. Though the cluster

16p11.2, distal
of chromatin or histone organization was only supported by

Locus (gene)
SCHEMA, the former two clusters (“channel or transporter

activities” and “neuronal components”) showed convergent
Table 1.
7q11.23
1q21.1
3q29
enrichment in both studies. Taken together, these can be

†
considered molecular pathways whose involvement in
5
Fig. 1 Allelic spectrum of schizophrenia-associated variants. A plot of minor allele frequencies in controls (x-axis) and ORs (y-axis) for
schizophrenia-associated genes/variants with robust statistical evidence, that is, the index SNPs of 287 genome-wide significant loci identified
by the PGC GWAS (blue, ref. [20]), ten exome-wide significant genes in the SCHEMA exome sequencing study (green, ref. [21]) and the study-
wide significant eight CNVs in the PGC CNV Working Groups study (red, ref. [31]). Genes and variants with OR > 1.2 were labeled. The sizes of
points are proportional to the −log10 P values for the association. The error bars indicate 95% confidence intervals of the ORs. The upper right
chronology summarizes the representative studies of etiologically valid mouse and cellular models of schizophrenia shown in Tables 2 and 3.
schizophrenia pathogenesis is supported by both common and of which five, including the top two (“TEGLU4: Excitatory
rare variant studies. neurons, Cortex pyramidal layer 5, Cingulate/Retrosplenial area
In addition to molecular pathways, both the PGC GWAS and (superficial and deep)” [1st in PGC GWAS and 7th in SCHEMA)
the SCHEMA study performed cell-type enrichment analysis and “TEGLU20: Excitatory neurons, Cortex pyramidal layer 6” [7th
utilizing data from single-cell RNA sequencing, which is rapidly in PGC GWAS and 4th in SCHEMA]), were annotated as deep layer
developing in recent years. For this analysis, both studies excitatory neurons. These results represent another form of
employed the statistical method described by Skene et al. [35] functional convergence of the findings from genetic studies of
and used the data of 265 cell types defined in a single-cell RNA common and rare variants, and provide insight into the cell types
sequencing study of the mouse nervous system by Zeisel et al. likely playing a critical role in the molecular pathology of
[36]. When we plotted the enrichment ranks for the 265 cell schizophrenia.
types in the PGC GWAS and the SCHEMA study, which are
detailed in Supplementary Fig. 3 and Supplementary Table 10 of Other types of variants potentially explain still-missing
the corresponding papers, respectively, we again found that heritability
there is a highly significant correlation (Fig. 2d, Pearson’s Although large-scale genetic studies and refinements in statistical
correlation coefficient = 0.74, P = 4.50 × 10−47) (note that we methods have elucidated a substantial part of the genetic
used the enrichment ranks because exact statistics were not architecture of schizophrenia, there remains a large gap between
available in Supplementary Fig. 3 of the PGC GWAS). Overall, the overall heritability reported in epidemiological studies
enrichment was particularly strong in excitatory neurons, (60–80%) [5–8] and that explained by common SNPs (24% in
followed by inhibitory neurons, and less pronounced in other ref. [20]) or rare gene-disruptive rare SNVs, indels, and CNVs (<10%
cell types such as glial, vascular, and immune cells. Among the according to refs. [37, 38]). Provisional evidence suggests other
top ten highest-ranked cell types, eight were excitatory neurons, types of variants that are not captured by GWAS or exome
6
sequencing are likely to be involved, including rare non-coding heritability. More detailed information on results from these
variants [39] and tandem repeat variants [40] identified through pioneering but preliminary studies can be found in Supplementary
whole genome sequencing. Also, there is preliminary evidence Note. To more accurately estimate the contributions from these
suggesting the role of postzygotic somatic variants [31, 41–44], under-studied types of variants, further large-scale studies are
while these are not transmitted and do not contribute to mandatory.
7
Fig. 2 Functional convergence of the findings from studies of common and rare variants. a A Venn diagram showing the overlap of GO
terms that showed enrichment (uncorrected P < 0.05) in the PGC GWAS and the SCHEMA exome sequencing study. b A plot of the enrichment
of 1431 GO terms commonly analyzed in the PGC GWAS and the SCHEMA study. The x- and y-axes indicate −log10 uncorrected P values in the
PGC GWAS and the SCHEMA study, respectively. The blue dotted lines indicate Bonferroni- or Benjamini-Hochberg-corrected significance
thresholds. GO terms with Benjamini-Hochberg-corrected P < 0.05 in both studies are indicated by labeled red dots, of which four terms with
Bonferroni-corrected P < 0.05 in both studies (voltage-gated cation channel activity, voltage-gated channel activity, voltage-gated ion channel
activity, and chemical synaptic transmission) are labeled in red. The correlation between the two studies (Pearson’s r = 0.39. P = 6.67 × 10−54) is
shown in the upper right. c Network visualization of the GO terms enriched in the PGC GWAS and the SCHEMA study. Significant GO terms
after the Bonferroni correction in either or both studies are displayed. Nodes of significant GO terms are color-coded as follows: pink, the PGC
GWAS; yellow, the SCHEMA study; red, both studies. The sizes of nodes are proportional to –log10 meta-analysis P values calculated by
combining uncorrected P values in the PGC GWAS and the SCHEMA study using Fisher’s method. Nodes are connected when the similarity
score ≥0.4, and the edge width is proportional to the similarity score. We observed three major clusters of GO terms, each related to channel
or transporter activities; neuronal components (synapse, axon, and dendrite); chromatin or histone organization. d A plot of the results of cell
type enrichment analyses in the PGC GWAS and the SCHEMA study. In both studies, the data of 265 cell types defined in the Zeisel et al. study
[36] were used. The x- and y-axes indicate enrichment ranks in the PGC GWAS and the SCHEMA study, respectively. The top ten cell types for
which the sum of the ranks in the PGC GWAS and the SCHEMA study is the smallest are labeled with the cell cluster ID, major cell type, and
likely location. Each circle indicates each cell type, which is color-coded as indicated in the lower right. The sizes of the circles are proportional
to the enrichment ranks. The correlation between the two studies (Pearson’s r = 0.74. P = 4.50 × 10−47) is shown in the upper left.
TRANSCRIPTOMIC AND EPIGENOMIC PATHOLOGY IN CRDs hyper-acetylated in schizophrenia, suggesting that dysregu-
SCHIZOPHRENIA lated H3K27ac peaks within dysregulated CRDs particularly are
GO enrichment analysis of rare coding variants identified associated with the genetic schizophrenia risk.
regulation of transcription and chromatin organization as one of Besides histone modifications, DNA methylation is another
the molecular pathways implicated in schizophrenia (Fig. 2c). Also, major epigenetic modifier with regulatory functions. Several
multiple single genes with large effect sizes, such as SP4 encoding studies have explored genome-wide DNA methylation status in
the transcription factor Sp4 and SETD1A encoding a histone postmortem schizophrenia brains. Among them, a study of
methyltransferase, are core components of transcriptional and microarray-based analysis of DNA methylation at ~450,000 loci
epigenetic regulation. In line with these findings, studies of in postmortem dorsolateral PFC (DLPFC) tissues from 526
transcriptomic and epigenomic pathology of schizophrenia using individuals was reported [50]. In this study, a total of 2104 sites
patient-derived tissues have been conducted at scale. differentially methylated between quality-controlled 108 schizo-
One of the largest studies of transcriptomic brain pathology in phrenia cases and 136 controls (Bonferroni-corrected P < 0.05), of
schizophrenia was conducted by the PsychENCODE consortium which 97.1% were hypomethylated in schizophrenia, were
[45]. In this study, gene- and transcript isoform-level differential identified. A GO enrichment analysis of genes in or near the
expression was comprehensively analyzed by performing RNA- differentially methylated sites showed an overrepresentation of
sequencing (RNA-seq) of bulk postmortem cerebral cortex tissues genes related to embryo development, cell fate commitment, and
from 559 schizophrenia cases and 936 control individuals, nervous system differentiation. Also, modest enrichment of
together with 51 ASD and 222 bipolar disorder brains. They schizophrenia-associated loci among the differentially methylated
identified that the expression of 4821 genes and 3803 isoforms sites (1.9% among differentially methylated sites vs. 1.3% among
significantly differed between schizophrenia and controls (FDR < others, P = 0.004, Chi-square test) was observed. On the other
0.05). Genes related to “inflammatory response” and “receptor hand, in a study of postmortem brain samples overlapping with
activity” were enriched in the significantly upregulated and the above-described microarray-based study (70 and 95 schizo-
downregulated genes/isoforms, respectively. The enrichment of phrenia DLPFC and hippocampus, and 77 and 102 control DLPFC
the genes related to receptor activities is consistent with the and hippocampus) using whole-genome bisulfite sequencing, a
results summarized in the previous section. Schizophrenia technique that can detect DNA methylation at the single base
heritability was enriched among genes and transcripts dysregu- resolution, much smaller numbers of differentially methylated
lated in schizophrenia brains, especially in down-regulated sites, none in DLPFC and 70 in the hippocampus, were identified
transcript isoforms. despite less stringent multiple testing corrections (FDR < 0.05) [51].
Regarding epigenomic brain pathology, a recent large-scale This discrepancy could be explained by the difference in sample
study analyzed two major histone modifications, histone 3 lysine sizes as well as the sensitivity to detect differentially methylated
27 acetylations (H3K27ac) and histone 3 lysine 4 trimethylations sites and the number of hypotheses tested, as a larger number of
(H3K4me3) [46], in postmortem prefrontal cortical samples (sorted sites are covered by whole-genome bisulfite sequencing.
neurons or bulk tissues) from 303 schizophrenia cases and 388 In addition to the studies using postmortem brain tissues, there
controls along with 48 bipolar disorder brains by chromatin are large-scale studies of peripheral samples aiming to identify
immunoprecipitation sequencing (ChIP-seq) [47]. In the analysis of disease biomarkers. In a study by Aberg et al. [52], analyzing
differential H3K4me3/H3K27ac peaks, 6219 differential H3K27ac genome-wide DNA methylation profiles in blood samples from
peaks (FDR < 0.05) between schizophrenia and controls were 759 schizophrenia cases and 738 controls by methyl-CpG–binding
identified though there were no differential H3K4me3 peaks. Of domain protein-enriched genome sequencing, 25 and 139 sites
these, schizophrenia heritability based on GWAS [20] was enriched associated with the diagnosis at Bonferroni-corrected P < 0.05 and
in the H3K27ac peaks hyper-acetylated in schizophrenia. Subse- FDR < 0.01, respectively, were identified. The most significant
quently, this study mapped cis-regulatory domains (CRDs), which association was observed in the region involving FAM63B, a part of
often overlap with topologically associating domains defined by networks regulated by microRNAs associated with neuronal
the analysis of 3D chromosomal conformations but constitute differentiation and dopaminergic gene expression. This associa-
smaller regulatory units of 104–106 bp [48, 49], in the brain using tion was replicated in an independent cohort of >1000 individuals.
the information of inter-individual correlations between histone The observed effect sizes for three associated methylation sites at
peaks. In an analysis integrating information on CRDs and FAM63B were moderate (Cohen’s d = 0.42–0.45). In a recent meta-
differential H3K27ac peaks, it was shown that schizophrenia analysis of blood DNA methylation profiles from 4483 participants
heritability is strongly enriched at differential H3K27ac peaks in from seven cohorts, including 1681 schizophrenia cases and 1583
8
controls, by Hannon et al. [53], 1013 differentially methylated loci spine morphology in Setd1a heterozygous knockout mice and
with methylome-wide significance (P < 9 × 10−8), which were several mouse models with CNVs [54, 55, 57, 58, 88, 91]. Common
annotated to 692 genes, were identified. Among 158 electrophysiological phenotypes include altered synaptic trans-
schizophrenia-associated loci identified by GWAS [10], overall missions, such as diminished excitability indicated by reduced
differential DNA methylation was observed at 21 loci after excitatory postsynaptic currents [55, 58, 60, 75, 79, 82, 85, 88, 91]
correcting for multiple testing, supporting co-localization of or deficits in long-term potentiation [68, 79], though some mice
signals from GWAS and epigenome-wide association study. On showed increased excitability or altered activities of other
the other hand, an integrative analysis of DNA methylation and neuronal subtypes such as GABAergic neurons [70, 84].
genetic variants exploring the causal relationships was not Behavioral alterations common in these mice include deficits in
performed in their study. Further studies of the interaction sociality, cognitive performance, and prepulse inhibition
between genetic and epigenetic factors that are expected to [54, 61, 66–68, 70–73, 75–77, 79, 88, 90–92]. These phenotypes
provide additional insights into the molecular mechanisms are generally consistent with those in human schizophrenia
underlying co-localization are warranted. patients [93], though there would be biases that these phenotypes
Overall, while the significant overlap between differentially are more likely to be investigated and reported. Also, in some
expressed or modified genes and the genetic risk of schizophrenia cases, there were inconsistent results even among models with
has been reported in some studies, transcriptomic or epigenomic mutations in the same gene. This could be explained by
alterations of single genes that can biologically define the general differences in the method of introduction of the mutations (e.g.,
population of schizophrenia or serve as a high-sensitivity and CRISPR/Cas9 or gene targeting), genetic backgrounds (e.g., C57BL/
specificity biomarker have not been discovered, or perhaps there 6J and C57BL/6N), and other factors. In addition, the acquisition of
is no such universal molecular marker. Therefore, further studies more definitive results will be facilitated by strict standardization
are necessary to identify conclusive transcriptomic and epige- of analysis protocols and ensuring a sufficient sample size, as have
nomic pathology in schizophrenia. been done in human genetics studies.
Cellular models
STUDIES OF ETIOLOGICALLY VALID MOUSE AND CELLULAR Recent technological advances have enabled the reproduction of
MODELS OF SCHIZOPHRENIA pathological conditions in vitro by creating patient-derived or
As summarized in Table 1 and Fig. 1, recent large-scale genetic mutation-carrying induced pluripotent stem (iPS) cells and then
studies have identified specific genes, SNVs/indels, and CNVs that differentiating them into central nervous system cells or miniature
confer a substantial risk of schizophrenia. Based on this, rodents or brains. Studies of etiologically valid cellular models of schizo-
cells carrying the alleles equivalent to the above-described risk phrenia produced with this technology, including those with
variants identified in humans have been generated and analyzed. 22q11.2 deletion, 16p11.2 deletion/duplication, SETD1A LOF/PTV,
In this section, we overview studies of such etiologically valid, i.e., and NRXN LOF/PTV, have been conducted and reported [94–106]
having the same causal conditions as in human patients, models (Table 3).
of schizophrenia. (Tables 2 and 3), which have provided various In line with the findings in etiologically valid mouse models,
insights into the connection between genetic pathology and molecular profiling of these cellular models generally supports
pathological changes at the levels of molecules (e.g., transcripts dysregulation of genes related to neural transmission, especially
and proteins), cells, neural circuits, whole tissues, or individuals’ synaptic genes [95–97, 100, 101, 106], transcriptional regulators
behaviors. including microRNAs [94, 104], and schizophrenia-associated
genes discovered by human genetics studies [96, 97, 104]. Also,
Mouse models morphological alteration of soma and dendrite were common
We systematically surveyed studies of mice with mutant alleles except for iPS cell-derived neurons with NRXN1 deletion
orthologous to the variants listed in Table 1 with an observed OR [94, 98, 101, 102, 106]. Abnormal neural activities are identified
greater than ten. We found that there are studies on the following in multiple patient-derived or genetically engineered iPS cell-
variants: 22q11.2 deletion, 16p11.2 deletion/duplication, 3q29 derived neurons, however, the directions of the abnormalities are
deletion, 15q11.2–13.1 duplication, 2p16.3 (NRXN1) deletion, sometimes inconsistent across models manipulating different
GRIN2A LOF/PTV, GRIA3 LOF/PTV, and SETD1A LOF/PTV (Table 2) genes [97, 101–106]. Though it may be due to artifacts depending
[54–92]. While there are studies of mice with mutations in other on the differences in the experimental designs, another possibility
genes, such as RB1CC1 (also known as FIP200), the introduced is that imbalanced excitatory/inhibitory activities themselves [107],
alleles were not equivalent to ones in human patients, and/or the regardless of the direction of abnormality, are important in
mice were not analyzed in the context of neuroscience, and schizophrenia pathology. It is also worth noting that multiple
thereby not highlighted in this review. studies have investigated interventions to improve abnormal
Regarding the molecular phenotypes mainly analyzed by phenotypes observed in cellular models [96, 100]. Overall, the iPS
transcriptomic profiling, commonly dysregulated pathways cell technology is a powerful tool to analyze the molecular
include neural transmission and regulation of transcription pathology of schizophrenia using human samples, and further
[54, 63, 70, 88, 91, 92], in agreement with the findings from studies are warranted.
human genetics and genomics studies. Also, analyses utilizing
results of large-scale human genetics studies have reported Considerations on the model validity
enrichment of genetic risk for schizophrenia in genes differentially In the previous sections, we have defined etiologically valid
expressed in the models or molecular targets of the genetically models using the criteria that the modified gene showed a
modified genes [88, 91]. Meanwhile, mutant mice originally significant association with schizophrenia after stringent multiple
created not as a schizophrenia model but to elucidate gene testing correction and that the ORs observed in studies that found
function in the central nervous system, such as knockout mice for the association was greater than ten. However, we would like to
GRIN2A or GRIA3 encoding a glutamate receptor subunit, have not explicitly state that there is still uncertainty regarding the validity
been subjected to omics analysis. Molecular profiling of these of these models.
etiologically valid models may provide further convergent insights First, it should be noted that in general, there are wide ranges of
into the brain pathology of schizophrenia. confidence intervals for ORs for rare risk variants (Fig. 1 and
Morphological analysis of neuronal cells in these models has Table 1). Indeed, another large-scale study (20,403 cases and
reported reduced axonal and dendritic complexity and abnormal 26,628 controls) analyzing the association of schizophrenia and
Table 2. Etiologically valid mouse models of schizophrenia.
Risk Type of Gene editing Common mutant Zygosity Background Ref. First author Year Key molecular Key cellular/ Other findings Behavioral phenotypes
variant in modification strategy allele name number phenotypes circuity
human phenotypes Sociality Cognitive ASR PPI Other
function behaviors
22q11.2 1.3 Mb Gene- Df(16)A+/- Heterozygous C57BL/6J 54 Stark KL. 2008 • Altered miRNA • Decreased Impaired Normal Low Hyperactive,
deletion deletion of targeting expression width of anxiety in
chr16 explained by Dgcr8 mushroom male mice
haploinsufficiency spines and
• Altered expression reduced
of "oxidative dendritic
phosphorylation" complexity in
and "ATP-synthesis- heterozygous
coupled electron knockout mice
transport" genes in of Dgcr8 within
PFC and 22q11.2
"Transmission of
nerve impulse" and
"Synaptic
transmission" genes
in the hippocampus
Heterozygous C57BL/6J 55 Mukai J. 2008 • Decreased • Rescue of the
density of morphological
dendritic spines, alteration by
glutamatergic restoration of
synapses, and Zdhhc8 in 22q11.2
dendritic
complexity
• Lower
miniature EPSC
frequency of
pyramidal
neuron
Heterozygous C57BL/6J 56 Sigurdsson T. 2010 • Reduced
hippocampal-
prefrontal
synchrony
Heterozygous C57BL/6J 57 Xu B. 2013 • Lower expression • Decreased
of miR-185 within spine density
22q11.2 and dendritic
• Higher expression complexity,
of Mirta22 (Emc10), a rescued by
target of miR-185 either
upregulation of
miR-185 or
downregulation
of Mirta22
Heterozygous C57BL/6J 58 Mukai J. 2015 • Disrupted • Altered
axonal growth, morphology of
branching, and neurons and
synaptic impaired
transmission prefrontal-
(smaller fEPSC in hippocampus
the frontal synchrony in

cortex) Zdhhc8+/- mice
Heterozygous C57BL/6J 59 Hamm JP. 2017 • Neuronal
activity with the
single-cell
resolution was
not altered, but
neuronal
coactivity
patterns
(ensembles)
were altered
around 1 Mb Gene- Df(16)1/+ Heterozygous C57BL/6J 60 Chun S. 2014 • Disruption of • Abnormal
deletion of targeting synaptic thalamocortical
chr16 transmission at sensitivity to
thalamocortical haloperidol
glutamatergic • Deficits of PPI in
projections in Dgcr8+/- mice
the auditory • Rescue of the
cortex caused deficiency of PPI in
by aberrant Dgcr8+/- mice by
elevation of haloperidol
Drd2 in the
thalamus
9
10
Table 2. continued
function behaviors
• Rescue effect
of an
antipsychotic,
haloperidol and
clozapine, on
the abnormal
thalamocortical
projection
3.0 Mb CRISPR/Cas9 Del(3.0Mb)/+ Heterozygous C57BL/6N 61 Saito R. 2020 • Decreased • Attenuation of Normal Impaired Increased Low Hypoactivity
deletion of expression of miR- visual-evoked
chr16 185-5p potential of the
primary visual
cortex (V1) and
the frontal
cortex
16p11.2 0.39 Mb Cre-mediated Del(7Slx1b-Sept1) Heterozygous C57BL/6N x 62 Horev G. 2011 • Generation of the Normal Altered
deletion deletion of recombination 4Aam 129Sv mice diurnal
chr7 • Enlargement of rhythms
multiple brain
structures
Heterozygous C57BL/6 x 63 Blumenthal I. 2014 • Altered
129Sv expressions
"regulation of
transcription" and
"regulation of actin
cytoskeleton
organization" genes
in the cortex
Heterozygous C57BL/6 x 64 Brunner D. 2015 • Growth deficits Normal Normal Normal Normal Normal USVs
129Sv
Heterozygous B6129SF1/J 65 Yin X. 2021 • Delayed spine Delayed

pruning motor
• Abnormally learning
high ensemble
activity of layer
II-III excitatory
neurons in the
motor cortex
during the initial
phase of
learning
0.44 Mb Cre-mediated Del(7Coro1a-Spn) Heterozygous C57BL/6N 66 Portmann T. 2014 • Altered • Increased • Generation of the Normal Impaired Decreased Hyperactivity
deletion of recombination 1Dolm dopaminergic number of mice
chr7 signaling in striatal medium • Growth deficits
neonatal brain spiny neurons • Anatomical
with dopamine abnormalities in
D2 receptor juvenile mice
(Drd2) and fewer
Drd1+ neurons
in deep layers of
cortex
• Abnormal
basal ganglia
circuitry
function
Heterozygous C57BL/6N 67 Yang M. 2015 • Growth deficits Reduced Decreased Low Fewer USVs,
anogenital (deafness) (deafness) reduced pain
sniffing sensitivity
0.46 Mb Cre-mediated Del(7Sult1a1-Spn) Heterozygous C57BL/6N 68 Arbogast T. 2016 • Underweight Normal Impaired Increased
deletion of recombination 6Yah despite human repetitive
chr7 carriers showing behaviors,
overweight increased
center time in
open field
16p11.2 0.39 Mb Cre-mediated Dp(7Slx1b-Sept1) Heterozygous C57BL/6N x 62 Horev G. 2011 • Generation of the Normal Altered
duplication duplication recombination 5Aam 129Sv mice diurnal
of chr7 rhythms,
increased
grooming
and resting
Table 2. continued
function behaviors
C57BL/6 x 63 Blumenthal I. 2014 • Altered
129Sv expressions of
"regulation of
transcription" and
"response to
protein stimulus"
genes in the cortex
C57BL/6 x 69 Blizinsky KD. 2016 • Identification of • Increased • Reversal of
129S7 Mapk3 within dendritic dendritic
16p11.2 as a central arborization phenotypes by
hub in MEK inhibitor
schizophrenia-
associated CNV
protein-protein
interaction network
C57BL/6 70 Rein B. 2021 • Upregulation of • Deficient • Rescue of Low Impaired Normal Normal Increased self-
enzyme modulator GABAergic synaptic and grooming,
genes and synaptic behavioral hypoactivity
downregulation of transmission phenotypes by
transcription factor and elevated restoration of
genes, including neuronal Npas4 in PFC
GABAergic synapse excitability in
regulator Npas4 in the prefrontal
PFC cortex
0.46 Mb Cre-mediated Dp(7Sult1a1-Spn) Heterozygous C57BL/6N 68 Arbogast T. 2016 • Smaller LTP in • Overweight Normal improved Hypoactivity,
duplication recombination 6Yah CA1 despite human decreased
of chr7 carriers showing center time in
underweight open field test
3q29 1.22 Mb Cre-mediated Heterozygous C57BL/6J 71 BaBa M. 2019 • Neuronal • Growth deficits, Low Impaired Increased Low Hyperactivity,
deletion deletion of recombination hyperactivation lower brain weight increased self-
chr16 and reduction of grooming
parvalbumin-
positive cells in
the cortex
1.26 Mb CRISPR/Cas9 Heterozygous C57BL/6N 72 Rutkowski TP. 2021 • Growth deficits Low Impaired Normal in Normal
deletion of • Partial but not in male male
chr16 complete Normal in Increased
reproduction of female in female
the phenotypes in
the deletion
model in mice
with
haploinsufficiency
of Dlg1 within
3q29
15q11.2- 6.3 Mb Cre-mediated Dp(7Herc2-Mkrn3) Heterozygous C57BL/6J or 73 Nakatani J. 2009 • Altered 5-HT2c Low Longer Increased Normal Fewer USV,
13.1 duplication recombination 1Taku 129SvEv receptor freezing in inflexibility,
duplication of chr7 signaling cued increased

condition anxiety,
of FC test depressive
behavior
2p16.3 Deletion of Gene- Nrxn1tm1Sud Homozygous SV129xC57BL/ 74 Geppert M. 1995 • Generation of
(NRXN1) exon1 targeting 6 Nrxn1α knockout
deletion mice
Homozygous SV129xC57BL/ 75 Etherton MR. 2009 • Reduced Normal Normal Increased Low Increased self-
6 spontaneous grooming,
excitatory impaired
synaptic nest-building
transmission in behaviors
CA1 pyramidal
neurons
• Decreased
evoked
excitatory
synaptic
strength in CA1
Homozygous C57BL/6J 76 Grayton HM. 2013 • No significant Aggressive Normal Impairment of
behavioral behavior locomotor
changes in Increased activity in
heterozygous KO social females,
mice interaction anxiety in
males,
11
12
Table 2. continued
function behaviors
impairment of
nest-building
behavior
Heterozygous SV129xC57BL/ 77 Dachtler J. 2015 Impaired Impaired Normal Normal
6 discrimination in female
Homozygous/ C57BL/6 78 Davatolhagh 2021 • Decreased • Altered
heterozygous MF. synaptic postsynaptic
strength NMDAR function
specifically onto at parafascicular-
indirect pathway DMS synapses
spiny projection (homozygous
neurons in only)
dorsal PFC-
dorsomedial
striatum (DMS)
circuit
(heterozygous
and
homozygous)
GRIN2A Disruption of Gene- Grin2atm1Mim Homozygous C57BL/6 x 79 Sakimura K. 1995 • Reduced • Generation of the Impaired
LOF/PTV the exon 10 targeting CBA channel current mice
(knockout) of NMDA
receptor and
hippocampal
LTP in CA1
pyramidal cells
Heterozygous C57BL/6 80 Takeuchi T. 2001 Normal Normal Homozygous
knockout
mice showed
increased
startle
responses
Heterozygous C57BL/6J 81 Marquardt K. 2014 Normal Homozygous
knockout
mice showed
cognitive
dysfunction in
set-shifting
task
Disruption of Gene- Grin2atm1Nak Homozygous 129/SvJ 82 Kadotani H. 1996 • Reduced • Generation of the Normal motor
the exon 10 targeting amplitude of the mice coordination
(knockout) slow component
of EPSC in
mossy fiber-
granule cells in
cerebellar slices
Heterozygous C57BL/6 83 Spooren W. 2004 • Impaired PPI by Normal Normal Homozygous
Inhibition of both knockout
GRNI2A/B mice did not
show any
significance in
PPI
GRIA3 LOF/ Disruption of Gene- Gria3tm1Zpj Hemizygous 129 x C57/BL6 84 Meng Y. 2003 • Normal basal • Generation of the No apparent
PTV exon 12 targeting synaptic mice behavioral
transmission deficits
• Normal (data not
presynaptic shown)
function and
LTD but
enhanced LTP
Hemizygous C57BL/6J 85 Peng SX. 2022 • Decreased • Improvement of Aggressive Reduced
excitation of aggression by behavior anxiety in the
mPFC neurons restoration of elevated plus
Gria3 in mPFC maze
Disruption of Gene- Gria3tm2Rlh Hemizygous C57BL/6J 86 Adamczyk A. 2012 • Increased Aggressive Normal Impaired
exon 12 targeting dopamine behavior motor
concentrations function,
in striatum and normal
reduced anxiety
serotonin
turnover in the
olfactory bulb
Table 2. continued
function behaviors
tm1Dgen
Disruption of Gene- Gria3 Hemizygous C57BL/6 87 Renner MC. 2017 • Impaired
exon 12 targeting cAMP-driven
postsynaptic
potentiation of
CA1 neurons
SETD1A Premature Gene- Setd1atm1a(EUCOMM) Heterozygous C57BL/6J 88 Mukai J. 2019 • Enrichment of • Decreased • Significant Normal Impaired
Wtsi
LOF/PTV stop codon targeting high-confidence mushroom overlap between
cassette target genes of spine density Setd1a and Mef2
in intron 3 Setd1a for synaptic and axonal targets at
and established branching but enhancers
schizophrenia/NDD- not dendritic
associated genes complexity in
PFC neurons
• Altered
neuronal short-
term plasticity
and excitability
in PFC
Heterozygous C57BL/6J 89 Hamm JP 2020 • Aberrant
cortical cell-cell
ensembles in V1
during ongoing
and visual
evoked activities
• Altered cortical
oscillations
Heterozygous C57BL/6J 90 Bosworth 2021 Increased Low Increased
ML. time spent in
inner zone in
open field test
2 bp CRISPR/Cas9 Heterozygous C57BL/6N 91 Nagahama K. 2020 • Enrichment of • Attenuated Low Impaired Normal Low Hypoactivity
deletion in DEGs in mPFC for excitatory
exon 7 neural/synaptic/ synaptic
axonal genes and function and
schizophrenia/NDD- altered structure
associated genes such as smaller
number of
spines in L2/3
pyramidal
neurons of the
medial PFC
8 bp CRISPR/Cas9 Heterozygous C57BL/6N 92 Chen R. 2022 •Larger numbers of • Impaired • Reduced Normal Normal Increased Low Lower sucrose
deletion in DEGs in Foxp2(+) exocytosis H3K4me3 in Foxp2 preference
exon 15 deep layer cortical functions (+) neurons
neurons and striatal • Decreased
D1- or D2-type spine density
medium spiny and dendritic
neurons complexity in
• Enrichment of PFC and striatal

synaptic genes in neurons
PFC excitatory
neuron DEGs
ASR acoustic startle response, DA dopamine, DEG differentially expressed gene, EPSC excitatory postsynaptic current, FC fear conditioning, fEPSC field excitatory postsynaptic current, iPS induced pluripotent stem
cell, LOF loss-of-function, LTD long term depression, LTP long term potentiation, mPFC medial PFC, NDD neurodevelopmental disorders, NPC neural progenitor cell, PFC prefrontal cortex, PPI prepulse inhibition,
PTV protein-truncating variant, USV ultrasonic vocalization.
13
14
Table 3. Etiologically valid cellular models of schizophrenia.
Risk Model type Differentiation Zygosity N (Case) N (Control) Ref. number First author Year Molecular Cellular/circuity Other findings
variant phenotypes phenotypes
in human
22q11.2 Patient-derived Neurosphere Heterozygous 2 4 94 Toyoshima M. 2016 • Downregulation of • Reduction of
deletion miR-17/92 and miR- neurosphere size,
106a/b clusters neural
differentiation,
neurite outgrowth,
cellular migration,
and neurogenic-to-
gliogenic
competence ratio
Patient-derived NPC, 2D- Heterozygous 8 7 95 Lin M. 2016 • Upregulation of • Lower NPC-
neurons genes related to proliferation rates
apoptosis/
programmed cell
death and
downregulation of
synaptic, cell cycle,
and microtubule
organization genes
Patient-derived Cerebral Heterozygous 15 15 96 Khan TA. 2020 • Altered expressions • Deficits of • Recapitulation of
cortical of neuronal neuronal calcium and
organoids excitability- excitability and membrane potential
related genes calcium signaling alterations in DGCR8+/-
• Enrichment of neurons
schizophrenia • Rescue by DGCR8
heritability in DEGs of restoration or an
mature organoids antipsychotic,
raclopride
Patient-derived iPS, NPC, 2D- Heterozygous 20 29 97 Nehme R. 2022 • Enrichment of • Lower spiking rate
excitatory schizophrenia of neurons in multi-
neurons genetic risk and electrode array
synaptic genes in analysis
upregulated DEGs of
NPCs and neurons
16p11.2 Patient-derived 2D-cortical Heterozygous 3 deletion 4 98 Deshpande A. 2017 • Increased and
deletion/ neurons 3 duplication decreased soma
duplication area/dendritic
length in 16p11.2
deletion and
duplication,
respectively
• Reduction of
synaptic density in
both deletion and
duplication
Patient-derived Cortical Heterozygous 6 deletion 4 99 Kostic M. 2021 • A larger number of • Similar cell
organoids 8 duplication DEGs in 16p11.2 composition across
deletion than in organoids derived
duplication from 16p11.2 CNV
• Downregulation of carriers (deletion or
neuron projection duplication) and
morphology-related controls as
genes and revealed by scRNA-
upregulation of seq
positive chemotaxis
genes in the
deletion lines
• Perturbation of
genes regulated by
RBFOX1 in the
deletion lines
Patient-derived Cortical Heterozygous 3 deletion 3 100 Urresti J. 2021 • Enrichment of • Accelerated • Rescue of migration
organoids 3 duplication “ligand-gated ion neural maturation deficits by inhibition
channel activity” in 16p11.2 deletion of RhoA activity
Table 3. continued
Risk Model type Differentiation Zygosity N (Case) N (Control) Ref. number First author Year Molecular Cellular/circuity Other findings
variant phenotypes phenotypes
in human
genes in DEGs of than in duplication
16p11.2 deletion • Increased synaptic
• No enrichment of puncta in 16p11.2
GO terms in DEGs of deletion
16p11.2 duplication • Neuronal
migration deficits
in both 16p11.2
deletion and
duplication
CRISPR/Cas9 2D- Heterozygous 3 deletion 3 101 Sundberg M. 2021 • Increased synaptic • Increased soma • Hyperactive DA
dopaminergic 3 duplication marker expression in size in 16p11.2 neuronal networks
neurons 16p11.2 deletion and increased
deletion lines • Deficits in bursting in 16p11.2
• Reduced synaptic neuronal deletion
marker expression in differentiation in
16p11.2 16p11.2 deletion
duplication lines
CRISPR/Cas9 2D-neurons Heterozygous 7 deletion 6 clones 102 Tai DJC. 2022 • Only nine DEGs in • Shortened • Altered proportion of
and cerebral 6 duplication (in the 2D- 16p11.2 deleted 2D- neurites with the cell components
organoids (in the 2D- neuron assay) neurons while 95 reduced of cerebral organoids
neuron assay) DEGs in duplicated branchpoints in in 16p11.2 deletion
neurons both 16p11.2 (i.e., decreased
• Enrichment of deleted and excitatory neurons
energy metabolism- duplicated 2D- and increased
related genes in neurons inhibitory GABAergic
DEGs of 16p11.2 • Reduced activity, neuron)
duplicated 2D- synchrony, and (2 clones per
neurons oscillation in 2D- genotype in the
neurons cerebral organoid
assays)
2p16.3 Deletion of exon 2D-excitatory Heterozygous 1 (exon 19 1 (exon 19 103 Pak C. 2015 • Normal neuronal
(NRXN1) 19 shared by all cortical deletion) deletion) morphology
deletion neurexin-1 isoforms neurons 2 (premature 2 (premature • Decreased
or insertion of stop codon) stop codon) spontaneous
premature stop mEPSC frequency
codon just before • Iimpaired
the last exon by Cre- neurotransmitter
loxP/FLP-FRT in release
human ES cells
Patient-derived 2D-neurons Heterozygous 2 (5’- 4 (including a 104 Flaherty E. 2019 • Enrichment of • Reduced • Comprehensive
NRXN1a+/-) family genes related to spontaneous cataloging of a total of
2 (NRXN1a/ member of transcriptional/ neuronal activity in 123 high-confidence
b+/-) NRXN1α/β+/- epigenetic regulation multi-electrode in-frame human

carrier) and schizophrenia array analysis NRXN1α isoforms
GWAS-associated
genes in DEGs of
patient-derived NPC
and neurons
Patient-derived 2D-neurons Heterozygous 3 carrier- 3 non-carrier- 105 Pak C. 2021 • No consistent • No change in
resembling derived derived differences between dendritic and
cortical layer 2/ carrier- and non- synaptic
3 pyramidal carrier-derived morphologies
neurons neurons in principle • Decrease in the
component analyses frequency of
of the data of bulk- spontaneous
RNA-seq excitatory synaptic
• Upregulation of events, in evoked
intracellular NRXN1- excitatory synaptic
binding protein transmission, and
CASK in patient- in paired-pulse
derived iPS-neurons depression
15
16
CNVs implicated in neurodevelopmental disorders reported no
statistically significant associations of 16p11.2 deletion and
15q11.2–13.1 duplication with schizophrenia and association of
Other findings
2p16.3 (NRXN1) deletion with OR smaller than ten [30, 32].

Therefore, there is the possibility that ORs are overestimated in the
existing data. Second, as mentioned in the section on genetic
pathology, many of the genes and variants highlighted here,
especially almost all CNVs, are associated not only with schizo-
phrenia but also with other neurodevelopmental disorders. Given
and arborization,
dendritic length
Cellular/circuity
this, the mice and cells harboring such mutations should not be
network burst
• Increase of
phenotypes
activity, and
considered as specifically modeling schizophrenia. Third, particu-

integration
DEG differentially expressed gene, GWAS genome-wide association study, iPS induced pluripotent stem cell, NPC neural progenitor cells, scRNA-seq single-cell RNA sequencing.
synaptic
larly in the case of CNV-based animal models, evaluation of the

model validity and the interpretation of the phenotypes require
caution because CNVs usually contain multiple genes and non-
coding regions whose structure is sometimes not well conserved
gene sets associated
between rodents and humans. Lastly, we would like to emphasize

with glutamatergic
• Increased cAMP
synaptic function
that a number of valuable findings interpreted as being relevant

• Perturbation in
to the molecular pathology of schizophrenia have also been

phenotypes
Molecular
obtained from analyses of models in which genes not strongly

signaling
supported by currently available evidence from human genetics

studies were manipulated. For example, given the mechanisms of
action of known antipsychotics, it is quite obvious that
2022
dysregulation of the monoaminergic system is involved in the

Year
pathophysiology of schizophrenia [108, 109], and therefore

various genes in this pathway have been intensively investigated.
Although these include genes that do not have strong genetic
First author
support, unlike DRD2 identified in GWAS [9, 20] and others, they
Wang S.
form a foundation not only for the study of schizophrenia patients

but also for the study of animal and cellular models with genetic
etiological validity [60, 110–114]. Besides, as evidenced by the fact
that heritability is better explained by considering numerous SNPs,
Ref. number
not only those in genome-wide significant loci, it is clear that there

are true schizophrenia risk genes among those that did not reach
106
the stringent significance threshold with the current sample sizes.

This indicates that further identification of robust risk genes in
larger studies will certainly increase the value of existing research
N (Control)
using animal and cellular models with modifications of such

1 clone
genes. Meanwhile, it is also true that there are mice that have
been interpreted as models of schizophrenia based on their face
validity, despite the lack of etiological validity in light of currently
available knowledge. Moreover, sometimes models are inter-
preted as meeting face validity based on phenotypes in
N (Case)
2 clones
homozygous mutants, even though heterozygous variants are

associated with schizophrenia in humans. Therefore, caution
should be exercised in discussing the validity of a model
solely on the strength of its face validity, regardless of the
Heterozygous
robustness of the association between the manipulated gene and

schizophrenia risk.
Zygosity
CURRENT ACHIEVEMENTS, LIMITATIONS, AND NEW

Differentiation
DIRECTIONS FOR FUTURE RESEARCH

2D-neurons
Based on the above-described existing knowledge of the genetic

and molecular pathology of schizophrenia as well as the emerging
insights into their links to other scales of pathologies from studies
of etiologically valid models, we summarize the current achieve-
ments, limitations, and new directions for future research as
follows.
frameshift deletion
Regarding the genetic pathology, significant advances, such as

(heterozygous
the elucidation of the highly polygenic nature of schizophrenia,

CRISPR/Cas9
Model type
in exon 7)
the explanation of more than 20% of disease liability by

continued
measurable genetic variants, and the identification of specific

genes and CNVs associated with schizophrenia with large effect
sizes, have been achieved. While a substantial part of the
heritability is still unexplained and the number of disease-
in human
LOF/PTV
Table 3.
variant
responsible genes identified so far is not as large as in ASD,

SETD1A
Risk
where similarly large studies have been conducted, it is certainly

expected that by simply increasing the sample size and
17
investigating under-studied types of variants, such as rare non- GWAS (and supporting evidence was obtained for 11 of the
coding mutations, we can better explain the genetic liability to remaining 12 loci in an extended analysis). Given these, it is
schizophrenia and identify additional responsible genes. Indeed, a warranted to further increase the sample sizes in postmortem
more recent target sequencing study of candidate schizophrenia- brain studies of schizophrenia in order to obtain robust and
associated genes in 11,580 cases and 10,555 controls, followed by reproducible results, as has been done in GWAS throughout its
a meta-analysis with the SCHEMA dataset identified two novel history. The same would be true for human brain imaging studies
exome-wide significant genes, AKAP11 and SRRM2 [115], confirm- that explore structural and functional alterations associated with
ing the importance of increasing sample sizes. In addition to the phenotypes in a brain-wide manner [123].
promotion of basic genetic research, the application of genetic Third, as is true in any field of science, often important
information to clinical psychiatry is a subject of active discussion. unresolved problems, such as the mystery of the molecular brain
As an example, there are attempts to utilize polygenic risk scores pathology of schizophrenia, are addressed by the utilization of
(PRS) based on the profiles of common SNPs to predict clinical new technologies. Considering the major limitations of current
courses [116, 117], though further studies are needed. Also, the research on molecular pathology using patient or model brains,
identification of patients with Mendelian genetic disease among while the etiological validity of the model is undoubtedly
patients clinically diagnosed with schizophrenia based on important, in this context, there is an inherent concern that the
operationalized criteria and the optimization of their treatment molecular pathology in human schizophrenia patients may not be
based on genetic diagnosis is expected to be implemented in the adequately reproduced in rodent or cellular models, even when
near future. the exact same variants that are pathogenic in humans are
Compared to the knowledge of genetic pathology, our under- introduced. Perhaps this problem would be addressed by studying
standing of the molecular pathology of schizophrenia, such as species closer to humans, specifically non-human primates. Owing
transcriptomic and epigenomic alterations, is insufficient and no to recent innovations in genome editing technology, genetically
convincing single molecular markers that can biologically define engineered non-human primates carrying mutations that are
the schizophrenic brain have been identified. Nevertheless, pathogenic in humans, such as cynomolgus monkeys with
collectively interpreting the results of large-scale omics analyses mutations in MECP2 [124], the gene responsible for Rett syndrome,
of human postmortem brains and studies of models with high or SHANK3 [125], an ASD gene in the Phelan–McDermid syndrome
etiological validity, one might be able to argue that small critical region, and common marmosets with a mutation in PSEN1
transcriptional and/or epigenetic alterations of many [126] causal for familial Alzheimer’s disease, have been created
schizophrenia-associated genes and genes involved in neuronal and analyzed. While research using primates is much more time-
processes such as the formation and regulation of synapses would and cost-consuming than studies of mice, non-human primate
be the underlying molecular brain pathology of schizophrenia. To models of schizophrenia will be an excellent resource to fill the
obtain a clearer picture, the following directions would be gap between humans and rodents. Another major limitation is
considered. that while we can comprehensively analyze transcriptomic and
First, as summarized in Tables 2 and 3, multiple etiologically epigenomic profiles in postmortem brain tissues, such analysis in
valid models of schizophrenia have been generated and analyzed. the brain of living patients can not be performed. On the other
One of the next important steps will be to elucidate the alterations hand, recent technological advances have allowed us to quantify
that are commonly observed across them, and studies seeking this some key molecules involved in the regulation of synapses and
goal should be facilitated by investigating two or more models in histone modifications, such as the synaptic vesicle glycoprotein 2A
the controlled same experimental settings. This is because the [127], AMPA-type glutamate receptors [128], and histone deace-
results of studies of disease models are often confounded by tylases (HDACs) [129] in the living human brain. By expanding the
subtle differences in experimental design and conditions, such as repertoire of measurable molecules and scaling up studies, we will
apparatus, experimenter, mouse strain and genetic background, better understand molecular changes in the brain of living
and others. However, to our knowledge, there have been no schizophrenic patients.
publications reporting the results of the analysis of multiple Besides them, one of the most prominent new technologies
schizophrenia models with high etiological validity listed in that have become prevalent over the last decade is single-cell
Tables 2 and 3 under the same conditions. By conducting such sequencing techniques, whose usefulness was mentioned in the
studies of multiple models, which have already been done for ASD above section describing the convergent results of cell type
[118–121], it is expected that we can obtain convergent insight enrichment analysis in the PGC GWAS and SCHEMA study. Single-
into the pathological changes in schizophrenia. cell analyses are particularly powerful in studies of organs where
Second, while it must be recognized that collecting human different cell types are intermingled, such as the brain. By
postmortem brains on a large scale requires a great endeavor, performing cell type-resolved analysis using single-cell technol-
there is an open question of whether the sample sizes in studies ogy, it may be possible to more clearly capture molecular
to date are sufficient. Evaluating the inter-study reproducibility, pathology that was obscured in bulk tissues. At the end of this
which may help answer this question, it is true that the result of section, we highlight pioneering single-cell (nucleus) RNA
the analysis of genes differentially expressed between schizo- sequencing studies of postmortem schizophrenia brains, while
phrenia cases and controls in the aforementioned PsychEncode some of them have been posted to preprint servers and have not
study [45] is well correlated with that of a preceding study by the been published after peer review.
CommonMind Consortium (CMC) [122] (correlation coefficient
between the two studies for 687 genes with FDR < 0.05 in the CMC Single-nucleus RNA sequencing studies of postmortem
study = 0.799). On the other hand, of the 23 genes that showed schizophrenia brains
statistical significance after Bonferroni correction in the CMC study Technically, analysis of single “cells”, including cytoplasm, cannot
(uncorrected P < 3.04 × 10−6, 0.05 divided by the number of genes be currently performed in studies of frozen postmortem brain
analyzed, 16,423), only nine genes surpassed the same signifi- tissues; therefore, single-“nucleus” RNA sequencing (snRNA-seq)
cance threshold in the PsychENCODE study. This number is much studies of human schizophrenia brains have been conducted.
more than random expectation; however, this contrasts with the To our knowledge, there are four publications on snRNA-seq of
observation in GWAS that 116 out of the 128 loci genome-wide postmortem schizophrenia brains, including two preprints that
significantly (P < 5 × 10−8) associated with schizophrenia in the have not yet been peer-reviewed. Among them, the largest study
previous PGC GWAS in 2014 were replicated with the genome- by Ruzicka et al. analyzed 266,431 nuclei from 24 schizophrenia
wide significant association in the same local regions in 2022 patients and 293,589 nuclei from 24 controls using frontopolar
18
cortex (Brodmann area 10) samples [130]. In this study, 20 cell situation at that time as “the best of times, the worst of times for
types/states were annotated based on their transcriptional psychiatric disease” [134]. This was because, on the one hand, the
profiles, and it was shown that the majority of genes differentially development and deployment of long-awaited new DNA sequen-
expressed in schizophrenia occurred in the neuronal population. cing technology (i.e., next-generation sequencing) made it
The cell types with the strongest enrichment of schizophrenia- possible to conduct genome-wide exploration of highly penetrant
associated genes identified by GWAS among differentially rare variants on a population scale (the best of times), while on the
expressed genes include cortico-cortical projection neurons in other hand, many pharmaceutical companies withdrew from the
the deep layers V/VI, parvalbumin-positive basket interneurons, research and development of novel therapeutics due to their low
and excitatory neurons of a novel cell state enriched in the success rates (the worst of the times). A decade later, as predicted,
supragranular layers II/III. This novel type of supragranular several robust risk genes with large effect sizes have been
excitatory neurons was more abundant in schizophrenia than in identified for schizophrenia, and the first results of pioneering
controls, but was preferentially found in schizophrenia individuals studies using animal and cellular models created on the basis of
with less “transcriptional pathology score”, defined by overall the discovery of these genes are beginning to be harvested
schizophrenia-associated transcriptional dysregulation in each [88–92, 106]. Overall, it can be said that we have achieved the
individual. Based on this observation, the authors speculated that expected outcomes over the past ten years. Also, a number of
this population of excitatory neurons, named Ex-SZTR, might be powerful new technologies have been developed and implemen-
associated with “schizophrenia transcriptional resilience”. While ted during this period. The important thing is to continue this
further scrutinization through peer reviews is needed, this finding progress, and such effort will reverse the retreat from research and
may contribute to conceptual advances in the understanding of development by pharmaceutical companies and other investors,
the molecular/cellular pathology of schizophrenia. which was recognized a decade ago and persists today.
The observation that the majority of differentially expressed In this context, it would be meaningful to provide a clearer
genes are found in neuronal populations was also reported in picture of how the field of schizophrenia genetics and biology will
another study by Reiner et al., where 127,930 and 145,120 nuclei further develop. In our view, the overarching challenge for the
from DLPFC of 12 schizophrenia and 14 control individuals were next decade will be how we translate the findings in basic genetic
analyzed by snRNA-seq, respectively [131]. In their study, ~96% of and biological research into clinical psychiatry. The first part of
differentially expressed genes were observed in neuronal cell the path to resolving this problem has been clarified by the
types, including excitatory neurons across layers II-V and results of studies to date. Aggregating the existing knowledge,
parvalbumin-positive interneurons. we are able to identify diagnostic genomic variants (e.g.,
In a study by Batiuk et al., not only snRNA-seq of sorted neurons Pathogenic or Likely Pathogenic variants in the American College
from DLPFC of 9 schizophrenia patients and 14 controls (81,817 and of Medical Genetics and Genomics [ACMG] guidelines [135]) in
127,236 nuclei, respectively) but also follow-up immunohistochem- 1–6% of schizophrenia patients by comprehensively analyzing
istry, single-molecule fluorescence in situ hybridization, and spatial rare variants [136–139], and to extract a small proportion of the
transcriptomics analyses in an extended cohort were performed population with high genetic risk (e.g., OR > 5) utilizing the overall
[132]. Results of these analyses convergently suggested that profiles of common variants (i.e., PRS [140, 141]). On the other
transcriptional dysregulation and altered cellular composition within hand, to our knowledge, there are no genetic tests for
the upper cortical layer, involving both GABAergic interneurons and schizophrenia approved by the government and covered by
principal projection (excitatory in the cortex) neurons, might be a health insurance. Among several reasons for this situation, the
core substrate associated with the brain pathology of schizophrenia. primary one is that the clinical benefits gained from genetic
These results would collectively support that schizophrenia is testing are far less than the cost and potential side effects. More
primarily a disease of neuronal cells. On the other hand, a unique specifically, there are two major factors limiting the benefits: the
study focusing on cells constituting the blood-brain barrier (BBB) performance of risk prediction from genetic information is
based on the neurovascular hypothesis of schizophrenia was insufficient, and the results of genetic testing rarely lead to
conducted by Puvogel et al. [133]. In their study, a total of 178,009 changes in clinical actions. To improve the performance of
nuclei (NEUN and OLIG2 negative nuclei enriched for BBB cells and genetic risk prediction, as described above, it is indispensable to
NEUN positive and OLIG2 negative nuclei enriched for neuronal expand the sample size and investigate various types of variants,
cells) from postmortem midbrain tissues of 15 schizophrenia which include not only common SNPs, rare coding SNVs, and
patients and 14 controls were analyzed by snRNA-seq. The results CNVs but also non-coding rare variants, repeat element variants,
showed that there was no significant difference in the relative complex structural variants, somatic variants, and others, with
proportions of the major BBB cell types between schizophrenia sufficient statistical power. In particular, the variants that are not
and controls. A limited number of genes were differentially common but not extremely rare, which can fill the blank region in
expressed in schizophrenia (14 genes with log2 fold change > 0.3 Fig. 1, will be a major target in future research. Also, it is crucial to
and FDR < 0.05). These differentially expressed genes were conduct sufficiently large studies in diverse ethnic populations.
restricted to ependymal cells and pericytes, suggesting that BBB The importance of such studies is evident from the observation
cells are not broadly affected in schizophrenia. that the performance of PRS is greatly reduced when the ethnicity
Overall, many of the findings in these studies are detectable only of the individuals being scored is different from that of the data
when cell type-resolved analysis is performed, demonstrating the used to construct the prediction model [142, 143]. Regarding the
value of snRNA-seq. Nevertheless, some of the above-described improvement of clinical actionability, it is expected that the
results should be considered preliminary because half of the four generation and investigation of multiple etiologically valid
studies highlighted here have not been peer-reviewed yet. Also, the schizophrenia models, as featured in this review, will play an
numbers of individuals analyzed in these studies are not large, while important role. The realization of precision medicine, such as
the numbers of nuclei examined were huge. Therefore, it is necessary gene therapy, for specific genetic diseases frequently comorbid
to consider whether the sample size is sufficient. with schizophrenia (e.g., 22q11.2 deletion or SETD1A haploinsuf-
ficiency syndrome) leveraging the observations in studies of
these models might be the achievable goal within the next
PERSPECTIVES: A DECADE AFTER THE BEST OF TIMES, THE decade. And beyond that, by integrating the results of human
WORST OF TIMES FOR PSYCHIATRIC DISEASE genetics and model studies as well as other areas of research,
In 2012, Karayiorgou et al. on behalf of the Genetic and Neural such as human functional imaging and brain circuity, it should be
Complexity in Psychiatry 2011 Working Group described the aimed to define biologically homogeneous schizophrenia
19
subgroups and identify the optimal treatment and prevention 24. Amberger JS, Bocchini CA, Scott AF, Hamosh A. OMIM.org: leveraging knowledge
for them. across phenotype-gene relationships. Nucleic Acids Res. 2019;47:D1038–43.
The World Health Organization estimates that by 2030 mental 25. Tansey KE, Rees E, Linden DE, Ripke S, Chambert KD, Moran JL, et al. Common
disorders will be the leading cause of disease burden globally [144], alleles contribute to schizophrenia in CNV carriers. Mol Psychiatry.
2016;21:1085–9.
of which a significant part should be accounted for by schizophrenia
26. Niemi MEK, Martin HC, Rice DL, Gallone G, Gordon S, Kelemen M, et al. Common
due to its chronic and often treatment-resistant nature. Studies genetic variants contribute to risk of rare severe neurodevelopmental disorders.
toward the elucidation of the molecular pathology of schizophrenia, Nature. 2018;562:268–71.
which forms the foundation for essential therapeutics, are of great 27. Takata A, Nakashima M, Saitsu H, Mizuguchi T, Mitsuhashi S, Takahashi Y, et al.
social value. Therefore, continuous investments from academia, Comprehensive analysis of coding variants highlights genetic complexity in
government, industry, and citizens, along with appropriate ethical, developmental and epileptic encephalopathy. Nat Commun. 2019;10:2506.
legal, and social considerations, are warranted. 28. Davies RW, Fiksinski AM, Breetvelt EJ, Williams NM, Hooper SR, Monfeuga T,
et al. Using common genetic variation to examine phenotypic expression and
risk prediction in 22q11.2 deletion syndrome. Nat Med. 2020;26:1912–8.
REFERENCES 29. Kirov G, Pocklington AJ, Holmans P, Ivanov D, Ikeda M, Ruderfer D, et al. De novo
CNV analysis implicates specific abnormalities of postsynaptic signalling com-
1. Jones HB. Pathology. In: Wexler P. (eds). Information resources in toxicology
plexes in the pathogenesis of schizophrenia. Mol Psychiatry. 2012;17:142–53.
(Fourth Edition). Academic Press: San Diego, 2009, pp 357–63.
30. Rees E, Kendall K, Pardiñas AF, Legge SE, Pocklington A, Escott-Price V, et al.
2. Griesinger W. Die Pathologie und therapie der psychischen krankheiten. (Krab-
Analysis of Intellectual Disability Copy Number Variants for Association With
be:Stuttgart, 1861).
Schizophrenia. JAMA Psychiatry. 2016;73:963–9.
3. Kraepelin E. Psychiatrie: ein Lehrbuch für Studierende und Aerzte. (Verlag von
31. Marshall CR, Howrigan DP, Merico D, Thiruvahindrapuram B, Wu W, Greer DS,
Johann Ambrosius Barth:Leipzig, 1899).
et al. Contribution of copy number variants to schizophrenia from a genome-
4. Jauhar S, Johnstone M, McKenna PJ. Schizophrenia. Lancet. 2022;399:473–86.
wide study of 41,321 subjects. Nat Genet. 2017;49:27–35.
5. Owen MJ, Sawa A, Mortensen PB. Schizophrenia. Lancet. 2016;388:86–97.
32. Rees E, Kirov G. Copy number variation and neuropsychiatric illness. Curr Opin
6. Lichtenstein P, Yip BH, Bjork C, Pawitan Y, Cannon TD, Sullivan PF, et al. Com-
Genet Dev. 2021;68:57–63.
mon genetic determinants of schizophrenia and bipolar disorder in Swedish
33. Kraft P. Curses-winner’s and otherwise-in genetic epidemiology. Epidemiology.
families: a population-based study. Lancet. 2009;373:234–9.
2008;19:649–51.
7. Sullivan PF, Kendler KS, Neale MC. Schizophrenia as a complex trait: evidence
34. de Leeuw CA, Mooij JM, Heskes T, Posthuma D. MAGMA: generalized gene-set
from a meta-analysis of twin studies. Arch Gen Psychiatry. 2003;60:1187–92.
analysis of GWAS data. PLoS Comput Biol. 2015;11:e1004219.
8. Wray NR. Gottesman II. Using summary data from the danish national registers
35. Skene NG, Bryois J, Bakken TE, Breen G, Crowley JJ, Gaspar HA, et al. Genetic
to estimate heritabilities for schizophrenia, bipolar disorder, and major
identification of brain cell types underlying schizophrenia. Nat Genet.
depressive disorder. Front Genet. 2012;3:118.
2018;50:825–33.
9. Schizophrenia Working Group of the Psychiatric Genomics Consortium. Biological
36. Zeisel A, Muñoz-Manchado AB, Codeluppi S, Lönnerberg P, La Manno G, Juréus
insights from 108 schizophrenia-associated genetic loci. Nature 2014;511:421–7.
A, et al. Brain structure. Cell types in the mouse cortex and hippocampus
10. Pardiñas AF, Holmans P, Pocklington AJ, Escott-Price V, Ripke S, Carrera N, et al.
revealed by single-cell RNA-seq. Science. 2015;347:1138–42.
Common schizophrenia alleles are enriched in mutation-intolerant genes and in
37. Visscher PM, Goddard ME, Derks EM, Wray NR. Evidence-based psychiatric
regions under strong background selection. Nat Genet. 2018;50:381–9.
genetics, AKA the false dichotomy between common and rare variant
11. Stefansson H, Ophoff RA, Steinberg S, Andreassen OA, Cichon S, Rujescu D, et al.
hypotheses. Mol Psychiatry. 2012;17:474–85.
Common variants conferring risk of schizophrenia. Nature 2009;460:744–7.
38. Gaugler T, Klei L, Sanders SJ, Bodea CA, Goldberg AP, Lee AB, et al. Most genetic
12. International Schizophrenia Consortium, Purcell SM, Wray NR, Stone JL, Visscher
risk for autism resides with common variation. Nat Genet. 2014;46:881–5.
PM, O’Donovan MC, et al. Common polygenic variation contributes to risk of
39. Halvorsen M, Huh R, Oskolkov N, Wen J, Netotea S, Giusti-Rodriguez P, et al.
schizophrenia and bipolar disorder. Nature. 2009;460:748–52.
Increased burden of ultra-rare structural variants localizing to boundaries of
13. Xu B, Roos JL, Dexheimer P, Boone B, Plummer B, Levy S, et al. Exome
topologically associated domains in schizophrenia. Nat Commun. 2020;11:1842.
sequencing supports a de novo mutational paradigm for schizophrenia. Nat
40. Mojarad BA, Engchuan W, Trost B, Backstrom I, Yin Y, Thiruvahindrapuram B,
Genet. 2011;43:864–8.
et al. Genome-wide tandem repeat expansions contribute to schizophrenia risk.
14. Xu B, Ionita-Laza I, Roos JL, Boone B, Woodrick S, Sun Y, et al. De novo gene
Mol Psychiatry. 2022;27:3692–8.
mutations highlight patterns of genetic and neural complexity in schizophrenia.
41. Maury EA, Sherman MA, Genovese G, Gilgenast TG, Rajarajan P, Flaherty E, et al.
Nat Genet. 2012;44:1365–9.
Schizophrenia-associated somatic copy number variants from 12,834 cases
15. Purcell SM, Moran JL, Fromer M, Ruderfer D, Solovieff N, Roussos P, et al. A reveal contribution to risk and recurrent, isoform-specific NRXN1 disruptions.
polygenic burden of rare disruptive mutations in schizophrenia. Nature. medRxiv. 2022; https://doi.org/10.1101/2021.12.24.21268385.
2014;506:185–90. 42. Kim MH, Kim IB, Lee J, Cha DH, Park SM, Kim JH, et al. Low-level brain somatic
16. Fromer M, Pocklington AJ, Kavanagh DH, Williams HJ, Dwyer S, Gormley P, et al. mutations are implicated in schizophrenia. Biol Psychiatry. 2021;90:35–46.
De novo mutations in schizophrenia implicate synaptic networks. Nature. 43. Fullard JF, Charney AW, Voloudakis G, Uzilov AV, Haroutunian V, Roussos P.
2014;506:179–84. Assessment of somatic single-nucleotide variation in brain tissue of cases with
17. Genovese G, Fromer M, Stahl EA, Ruderfer DM, Chambert K, Landén M, et al. schizophrenia. Transl Psychiatry. 2019;9:21.
Increased burden of ultra-rare protein-altering variants among 4,877 individuals 44. Bae T, Fasching L, Wang Y, Shin JH, Suvakov M, Jang Y, et al. Analysis of somatic
with schizophrenia. Nat Neurosci. 2016;19:1433–41. mutations in 131 human brains reveals aging-associated hypermutability. Sci-
18. Singh T, Kurki MI, Curtis D, Purcell SM, Crooks L, McRae J, et al. Rare loss-of- ence. 2022;377:511–7.
function variants in SETD1A are associated with schizophrenia and develop- 45. Gandal MJ, Zhang P, Hadjimichael E, Walker RL, Chen C, Liu S, et al.
mental disorders. Nat Neurosci. 2016;19:571–7. Transcriptome-wide isoform-level dysregulation in ASD, schizophrenia, and
19. McCarthy SE, Gillis J, Kramer M, Lihm J, Yoon S, Berstein Y, et al. De novo mutations bipolar disorder. Science. 2018;362:6420.
in schizophrenia implicate chromatin remodeling and support a genetic overlap 46. Gates LA, Foulds CE, O’Malley BW. Histone marks in the ‘driver’s seat’: functional
with autism and intellectual disability. Mol Psychiatry. 2014;19:652–8. roles in steering the transcription cycle. Trends Biochem Sci. 2017;42:977–89.
20. Trubetskoy V, Pardiñas AF, Qi T, Panagiotaropoulou G, Awasthi S, Bigdeli TB, 47. Girdhar K, Hoffman GE, Bendl J, Rahman S, Dong P, Liao W, et al. Chromatin
et al. Mapping genomic loci implicates genes and synaptic biology in schizo- domain alterations linked to 3D genome organization in a large cohort of
phrenia. Nature. 2022;604:502–8. schizophrenia and bipolar disorder brains. Nat Neurosci. 2022;25:474–83.
21. Singh T, Poterba T, Curtis D, Akil H, Al Eissa M, Barchas JD, et al. Rare coding 48. Waszak SM, Delaneau O, Gschwind AR, Kilpinen H, Raghav SK, Witwicki RM, et al.
variants in ten genes confer substantial risk for schizophrenia. Nature. Population variation and genetic control of modular chromatin architecture in
2022;604:509–16. humans. Cell 2015;162:1039–50.
22. Benner C, Spencer CC, Havulinna AS, Salomaa V, Ripatti S, Pirinen M. FINEMAP: 49. Delaneau O, Zazhytska M, Borel C, Giannuzzi G, Rey G, Howald C, et al. Chro-
efficient variable selection using summary data from genome-wide association matin three-dimensional interactions mediate genetic effects on gene expres-
studies. Bioinformatics. 2016;32:1493–501. sion. Science. 2019;364:6439.
23. Meng XH, Chen XD, Greenbaum J, Zeng Q, You SL, Xiao HM, et al. Integration of 50. Jaffe AE, Gao Y, Deep-Soboslay A, Tao R, Hyde TM, Weinberger DR, et al.
summary data from GWAS and eQTL studies identified novel causal BMD genes Mapping DNA methylation across development, genotype and schizophrenia in
with functional predictions. Bone 2018;113:41–8. the human frontal cortex. Nat Neurosci. 2016;19:40–7.
20
51. Perzel Mandell KA, Eagles NJ, Wilton R, Price AJ, Semick SA, Collado-Torres L, 75. Etherton MR, Blaiss CA, Powell CM, Südhof TC. Mouse neurexin-1alpha deletion
et al. Genome-wide sequencing-based identification of methylation quantitative causes correlated electrophysiological and behavioral changes consistent with
trait loci and their role in schizophrenia risk. Nat Commun. 2021;12:5251. cognitive impairments. Proc Natl Acad Sci USA. 2009;106:17998–8003.
52. Aberg KA, McClay JL, Nerella S, Clark S, Kumar G, Chen W, et al. Methylome-Wide 76. Grayton HM, Missler M, Collier DA, Fernandes C. Altered social behaviours in
Association Study of Schizophrenia: Identifying Blood Biomarker Signatures of neurexin 1α knockout mice resemble core symptoms in neurodevelopmental
Environmental Insults. JAMA Psychiatry. 2014;71:255–64. disorders. PLoS One. 2013;8:e67114.
53. Hannon E, Dempster EL, Mansell G, Burrage J, Bass N, Bohlken MM, et al. DNA 77. Dachtler J, Ivorra JL, Rowland TE, Lever C, Rodgers RJ, Clapcote SJ. Heterozygous
methylation meta-analysis reveals cellular alterations in psychosis and markers deletion of α-neurexin I or α-neurexin II results in behaviors relevant to autism
of treatment-resistant schizophrenia. eLife. 2021;10:e58430. and schizophrenia. Behav Neurosci. 2015;129:765–76.
54. Stark KL, Xu B, Bagchi A, Lai WS, Liu H, Hsu R, et al. Altered brain microRNA 78. Davatolhagh MF, Fuccillo MV. Neurexin1α differentially regulates synaptic effi-
biogenesis contributes to phenotypic deficits in a 22q11-deletion mouse model. cacy within striatal circuits. Cell Rep. 2021;34:108773.
Nat Genet. 2008;40:751–60. 79. Sakimura K, Kutsuwada T, Ito I, Manabe T, Takayama C, Kushiya E, et al. Reduced
55. Mukai J, Dhilla A, Drew LJ, Stark KL, Cao L, MacDermott AB, et al. Palmitoylation- hippocampal LTP and spatial learning in mice lacking NMDA receptor
dependent neurodevelopmental deficits in a mouse model of 22q11 micro- ε1 subunit. Nature. 1995;373:151–5.
deletion. Nat Neurosci. 2008;11:1302–10. 80. Takeuchi T, Kiyama Y, Nakamura K, Tsujita M, Matsuda I, Mori H, et al. Roles of
56. Sigurdsson T, Stark KL, Karayiorgou M, Gogos JA, Gordon JA. Impaired the glutamate receptor ε2 and δ2 subunits in the potentiation and prepulse
hippocampal-prefrontal synchrony in a genetic mouse model of schizophrenia. inhibition of the acoustic startle reflex. Eur J Neurosci. 2001;14:153–60.
Nature. 2010;464:763–7. 81. Marquardt K, Saha M, Mishina M, Young JW, Brigman JL. Loss of GluN2A-
57. Xu B, Hsu PK, Stark KL, Karayiorgou M, Gogos JA. Derepression of a neuronal containing NMDA receptors impairs extra-dimensional set-shifting. Genes Brain
inhibitor due to miRNA dysregulation in a schizophrenia-related microdeletion. Behav. 2014;13:611–7.
Cell. 2013;152:262–75. 82. Kadotani H, Hirano T, Masugi M, Nakamura K, Nakao K, Katsuki M, et al. Motor
58. Mukai J, Tamura M, Fénelon K, Rosen AM, Spellman TJ, Kang R, et al. Molecular discoordination results from combined gene disruption of the NMDA receptor
substrates of altered axonal growth and brain connectivity in a mouse model of NR2A and NR2C subunits, but not from single disruption of the NR2A or NR2C
schizophrenia. Neuron. 2015;86:680–95. subunit. J Neurosci. 1996;16:7859–67.
59. Hamm JP, Peterka DS, Gogos JA, Yuste R. Altered cortical ensembles in mouse 83. Spooren W, Mombereau C, Maco M, Gill R, Kemp JA, Ozmen L, et al. Pharma-
models of schizophrenia. Neuron. 2017;94:153–67.e8. cological and genetic evidence indicates that combined inhibition of NR2A and
60. Chun S, Westmoreland JJ, Bayazitov IT, Eddins D, Pani AK, Smeyne RJ, et al. NR2B subunit-containing NMDA receptors is required to disrupt prepulse inhi-
Specific disruption of thalamic inputs to the auditory cortex in schizophrenia bition. Psychopharmacology. 2004;175:99–105.
models. Science. 2014;344:1178–82. 84. Meng Y, Zhang Y, Jia Z. Synaptic transmission and plasticity in the absence of
61. Saito R, Koebis M, Nagai T, Shimizu K, Liao J, Wulaer B, et al. Comprehensive AMPA glutamate receptor GluR2 and GluR3. Neuron. 2003;39:163–76.
analysis of a novel mouse model of the 22q11.2 deletion syndrome: a model 85. Peng SX, Pei J, Rinaldi B, Chen J, Ge YH, Jia M, et al. Dysfunction of AMPA
with the most common 3.0-Mb deletion at the human 22q11.2 locus. Transl receptor GluA3 is associated with aggressive behavior in human. Mol Psychiatry.
Psychiatry. 2020;10:35. 2022;27:4092–102.
62. Horev G, Ellegood J, Lerch JP, Son YE, Muthuswamy L, Vogel H, et al. Dosage- 86. Adamczyk A, Mejias R, Takamiya K, Yocum J, Krasnova IN, Calderon J, et al.
dependent phenotypes in models of 16p11.2 lesions found in autism. Proc Natl GluA3-deficiency in mice is associated with increased social and aggressive
Acad Sci USA. 2011;108:17076–81. behavior and elevated dopamine in striatum. Behav Brain Res. 2012;229:265–72.
63. Blumenthal I, Ragavendran A, Erdin S, Klei L, Sugathan A, Guide Jolene R, et al. 87. Renner MC, Albers EH, Gutierrez-Castellanos N, Reinders NR, van Huijstee AN,
Transcriptional consequences of 16p11.2 deletion and duplication in mouse Xiong H, et al. Synaptic plasticity through activation of GluA3-containing AMPA-
cortex and multiplex autism families. Am J Hum Genet. 2014;94:870–83. receptors. Elife. 2017;6:e25462.
64. Brunner D, Kabitzke P, He D, Cox K, Thiede L, Hanania T, et al. Comprehensive 88. Mukai J, Cannavò E, Crabtree GW, Sun Z, Diamantopoulou A, Thakur P, et al.
analysis of the 16p11.2 deletion and null Cntnap2 mouse models of autism Recapitulation and reversal of schizophrenia-related phenotypes in setd1a-
spectrum disorder. PLoS One. 2015;10:e0134572. deficient mice. Neuron 2019;104:471–87.e12.
65. Yin X, Jones N, Yang J, Asraoui N, Mathieu ME, Cai L, et al. Delayed motor 89. Hamm JP, Shymkiv Y, Mukai J, Gogos JA, Yuste R. Aberrant cortical ensembles
learning in a 16p11.2 deletion mouse model of autism is rescued by locus and schizophrenia-like sensory phenotypes in Setd1a(+/−) mice. Biol Psy-
coeruleus activation. Nat Neurosci. 2021;24:646–57. chiatry. 2020;88:215–23.
66. Portmann T, Yang M, Mao R, Panagiotakos G, Ellegood J, Dolen G, et al. Beha- 90. Bosworth ML, Isles AR, Wilkinson LS, Humby T. Behavioural consequences of
vioral abnormalities and circuit defects in the basal ganglia of a mouse model of Setd1a haploinsufficiency in mice: evidence for heightened emotional reactivity
16p11.2 deletion syndrome. Cell Rep. 2014;7:1077–92. and impaired sensorimotor gating. bioRxiv. 2021; https://doi.org/10.1101/
67. Yang M, Mahrt EJ, Lewis F, Foley G, Portmann T, Dolmetsch RE, et al. 16p11.2 2021.12.10.471949.
deletion syndrome mice display sensory and ultrasonic vocalization deficits 91. Nagahama K, Sakoori K, Watanabe T, Kishi Y, Kawaji K, Koebis M, et al.
during social interactions. Autism Res. 2015;8:507–21. Setd1a insufficiency in mice attenuates excitatory synaptic function and
68. Arbogast T, Ouagazzal AM, Chevalier C, Kopanitsa M, Afinowi N, Migliavacca E, recapitulates schizophrenia-related behavioral abnormalities. Cell Rep.
et al. Reciprocal effects on neurocognitive and metabolic phenotypes in mouse 2020;32:108126.
models of 16p11.2 deletion and duplication syndromes. PLoS Genet. 92. Chen R, Liu Y, Djekidel MN, Chen W, Bhattacherjee A, Chen Z, et al. Cell type-
2016;12:e1005709. specific mechanism of Setd1a heterozygosity in schizophrenia pathogenesis. Sci
69. Blizinsky KD, Diaz-Castro B, Forrest MP, Schürmann B, Bach AP, Martin-de- Adv. 2022;8:eabm1077.
Saavedra MD, et al. Reversal of dendritic phenotypes in 16p11.2 microduplica- 93. Mena A, Ruiz-Salas JC, Puentes A, Dorado I, Ruiz-Veguilla M, De la Casa LG.
tion mouse model neurons by pharmacological targeting of a network hub. Reduced prepulse inhibition as a biomarker of schizophrenia. Front Behav
Proc Natl Acad Sci USA. 2016;113:8520–5. Neurosci. 2016;10:202.
70. Rein B, Tan T, Yang F, Wang W, Williams J, Zhang F, et al. Reversal of synaptic 94. Toyoshima M, Akamatsu W, Okada Y, Ohnishi T, Balan S, Hisano Y, et al. Analysis
and behavioral deficits in a 16p11.2 duplication mouse model via restoration of of induced pluripotent stem cells carrying 22q11.2 deletion. Transl Psychiatry.
the GABA synapse regulator Npas4. Mol Psychiatry. 2021;26:1967–79. 2016;6:e934.
71. Baba M, Yokoyama K, Seiriki K, Naka Y, Matsumura K, Kondo M, et al. Psychiatric- 95. Lin M, Pedrosa E, Hrabovsky A, Chen J, Puliafito BR, Gilbert SR, et al. Integrative
disorder-related behavioral phenotypes and cortical hyperactivity in a mouse transcriptome network analysis of iPSC-derived neurons from schizophrenia and
model of 3q29 deletion syndrome. Neuropsychopharmacology. 2019;44: schizoaffective disorder patients with 22q11.2 deletion. BMC Syst Biol.
2125–35. 2016;10:105.
72. Rutkowski TP, Purcell RH, Pollak RM, Grewenow SM, Gafford GM, Malone T, et al. 96. Khan TA, Revah O, Gordon A, Yoon SJ, Krawisz AK, Goold C, et al. Neuronal
Behavioral changes and growth deficits in a CRISPR engineered mouse model of defects in a human cellular model of 22q11.2 deletion syndrome. Nat Med.
the schizophrenia-associated 3q29 deletion. Mol Psychiatry. 2021;26:772–83. 2020;26:1888–98.
73. Nakatani J, Tamada K, Hatanaka F, Ise S, Ohta H, Inoue K, et al. Abnormal 97. Nehme R, Pietiläinen O, Artomov M, Tegtmeyer M, Valakh V, Lehtonen L, et al.
behavior in a chromosome-engineered mouse model for human 15q11-13 The 22q11.2 region regulates presynaptic gene-products linked to schizo-
duplication seen in autism. Cell. 2009;137:1235–46. phrenia. Nat Commun. 2022;13:3690.
74. Geppert M, Khvotchev M, Krasnoperov V, Goda Y, Missler M, Hammer RE, et al. 98. Deshpande A, Yadav S, Dao DQ, Wu ZY, Hokanson KC, Cahill MK, et al. Cellular
Neurexin I alpha is a major alpha-latrotoxin receptor that cooperates in alpha- phenotypes in human iPSC-derived neurons from a genetic model of autism
latrotoxin action. J Biol Chem. 1998;273:1705–10. spectrum disorder. Cell Rep. 2017;21:2678–87.
21
99. Kostic M, Raymond JJ, Henry B, Tumkaya T, Khlghatyan J, Dvornik J, et al. Patient 124. Chen Y, Yu J, Niu Y, Qin D, Liu H, Li G, et al. Modeling Rett syndrome using
brain organoids identify a link between the 16p11.2 copy number variant and TALEN-edited MECP2 mutant cynomolgus monkeys. Cell 2017;169:945–55.e10.
the RBFOX1 gene. bioRxiv. 2021; https://doi.org/10.1101/2021.11.21.469432. 125. Zhou Y, Sharma J, Ke Q, Landman R, Yuan J, Chen H, et al. Atypical behaviour
100. Urresti J, Zhang P, Moran-Losada P, Yu NK, Negraes PD, Trujillo CA, et al. Cortical and connectivity in SHANK3-mutant macaques. Nature 2019;570:326–31.
organoids model early brain development disrupted by 16p11.2 copy number 126. Sato K, Sasaguri H, Kumita W, Inoue T, Kurotaki Y, Nagata K, et al. A non-human
variants in autism. Mol Psychiatry. 2021;26:7560–80. primate model of familial Alzheimer’s disease. bioRxiv. 2020; https://doi.org/
101. Sundberg M, Pinson H, Smith RS, Winden KD, Venugopal P, Tai DJC, et al. 10.1101/2020.08.24.264259.
16p11.2 deletion is associated with hyperactivation of human iPSC-derived 127. Finnema SJ, Nabulsi NB, Eid T, Detyniecki K, Lin SF, Chen MK, et al. Imaging
dopaminergic neuron networks and is rescued by RHOA inhibition in vitro. Nat synaptic density in the living human brain. Sci Transl Med. 2016;8:348ra96.
Commun. 2021;12:2897. 128. Miyazaki T, Nakajima W, Hatano M, Shibata Y, Kuroki Y, Arisawa T, et al. Visua-
102. Tai DJC, Razaz P, Erdin S, Gao D, Wang J, Nuttle X, et al. Tissue- and cell-type- lization of AMPA receptors in living human brain with positron emission
specific molecular and functional signatures of 16p11.2 reciprocal genomic tomography. Nat Med. 2020;26:281–8.
disorder across mouse brain and human neuronal models. Am J Hum Genet. 129. Wey HY, Gilbert TM, Zürcher NR, She A, Bhanot A, Taillon BD, et al. Insights into
2022;109:1789-813. neuroepigenetics through human histone deacetylase PET imaging. Sci Transl
103. Pak C, Danko T, Zhang Y, Aoto J, Anderson G, Maxeiner S, et al. Human neu- Med. 2016;8:351ra106.
ropsychiatric disease modeling using conditional deletion reveals synaptic 130. Ruzicka WB, Mohammadi S, Davila-Velderrain J, Subburaju S, Tso DR, Hourihan
transmission defects caused by heterozygous mutations in NRXN1. Cell Stem M, et al. Single-cell dissection of schizophrenia reveals neurodevelopmental-
Cell. 2015;17:316–28. synaptic axis and transcriptional resilience. medRxiv. 2020; https://doi.org/
104. Flaherty E, Zhu S, Barretto N, Cheng E, Deans PJM, Fernando MB, et al. Neuronal 10.1101/2020.11.06.20225342.
impact of patient-specific aberrant NRXN1α splicing. Nat Genet. 2019;51:1679–90. 131. Reiner BC, Crist RC, Stein LM, Weller AE, Doyle GA, Arauco-Shapiro G, et al. Single-
105. Pak C, Danko T, Mirabella VR, Wang J, Liu Y, Vangipuram M, et al. Cross-platform nuclei transcriptomics of schizophrenia prefrontal cortex primarily implicates
validation of neurotransmitter release impairments in schizophrenia patient- neuronal subtypes. bioRxiv. 2021; https://doi.org/10.1101/2020.07.29.227355.
derived NRXN1-mutant neurons. Proc Natl Acad Sci USA. 2021;118:e2025598118. 132. Batiuk MY, Tyler T, Dragicevic K, Mei S, Rydbirk R, Petukhov V, et al. Upper
106. Wang S, Rhijn JV, Akkouh I, Kogo N, Maas N, Bleeck A, et al. Loss-of-function cortical layer-driven network impairment in schizophrenia. Sci Adv.
variants in the schizophrenia risk gene SETD1A alter neuronal network activity 2022;8:eabn8367.
in human neurons through the cAMP/PKA pathway. Cell Rep. 2022;39:110790. 133. Puvogel S, Alsema A, Kracht L, Webster MJ, Weickert CS, Sommer IEC, et al.
107. Liu Y, Ouyang P, Zheng Y, Mi L, Zhao J, Ning Y, et al. A selective review of the Single-nucleus RNA sequencing of midbrain blood-brain barrier cells in schi-
excitatory-inhibitory imbalance in schizophrenia: underlying biology, genetics, zophrenia reveals subtle transcriptional changes with overall preservation of
microcircuits, and symptoms. Front Cell Dev Biol. 2021;9:664535. cellular proportions and phenotypes. Mol Psychiatry. 2022;27:4731–40.
108. Meltzer HY, Stahl SM. The dopamine hypothesis of schizophrenia: a review. 134. Karayiorgou M, Flint J, Gogos JA, Malenka RC. Genetic and neural complexity in
Schizophr Bull. 1976;2:19–76. psychiatry 2011 Working Group. The best of times, the worst of times for psy-
109. Barnes NM, Sharp T. A review of central 5-HT receptors and their function. chiatric disease. Nat Neurosci. 2012;15:811–2.
Neuropharmacology. 1999;38:1083–152. 135. Richards S, Aziz N, Bale S, Bick D, Das S, Gastier-Foster J, et al. Standards and
110. Drago J, Gerfen CR, Lachowicz JE, Steiner H, Hollon TR, Love PE, et al. Altered guidelines for the interpretation of sequence variants: a joint consensus
striatal function in a mutant mouse lacking D1A dopamine receptors. Proc Natl recommendation of the American College of Medical Genetics and Genomics
Acad Sci USA. 1994;91:12564–8. and the Association for Molecular Pathology. Genet Med. 2015;17:405–24.
111. Baik JH, Picetti R, Saiardi A, Thiriet G, Dierich A, Depaulis A, et al. Parkinsonian- 136. Alkelai A, Greenbaum L, Docherty AR, Shabalin AA, Povysil G, Malakar A, et al.
like locomotor impairment in mice lacking dopamine D2 receptors. Nature. The benefit of diagnostic whole genome sequencing in schizophrenia and other
1995;377:424–8. psychotic disorders. Mol Psychiatry. 2022;27:1435–47.
112. Giros B, Jaber M, Jones SR, Wightman RM, Caron MG. Hyperlocomotion and 137. Zoghbi AW, Dhindsa RS, Goldberg TE, Mehralizade A, Motelow JE, Wang X, et al.
indifference to cocaine and amphetamine in mice lacking the dopamine High-impact rare genetic variants in severe schizophrenia. Proc Natl Acad Sci
transporter. Nature. 1996;379:606–12. USA. 2021;118:e2112560118.
113. Ramboz S, Oosting R, Amara DA, Kung HF, Blier P, Mendelsohn M, et al. Ser- 138. Mojarad BA, Yin Y, Manshaei R, Backstrom I, Costain G, Heung T, et al. Genome
otonin receptor 1A knockout: an animal model of anxiety-related disorder. Proc sequencing broadens the range of contributing variants with clinical implica-
Natl Acad Sci USA. 1998;95:14476–81. tions in schizophrenia. Transl Psychiatry. 2021;11:84.
114. Bengel D, Murphy DL, Andrews AM, Wichems CH, Feltner D, Heils A, et al. 139. Balakrishna T, Curtis D. Assessment of potential clinical role for exome
Altered brain serotonin homeostasis and locomotor insensitivity to 3, sequencing in schizophrenia. Schizophr Bull. 2020;46:328–35.
4-methylenedioxymethamphetamine (“Ecstasy”) in serotonin transporter- 140. Murray GK, Lin T, Austin J, McGrath JJ, Hickie IB, Wray NR. Could polygenic risk
deficient mice. Mol Pharm. 1998;53:649–55. scores be useful in psychiatry? A review. JAMA Psychiatry. 2021;78:210–9.
115. Liu D, Meyer D, Fennessy B, Feng C, Cheng E, Johnson JS, et al. Rare schizo- 141. Khera AV, Chaffin M, Aragam KG, Haas ME, Roselli C, Choi SH, et al. Genome-
phrenia risk variant burden is conserved in diverse human populations. wide polygenic scores for common diseases identify individuals with risk
medRxiv. 2022;https://doi.org/10.1101/2022.01.03.22268662. equivalent to monogenic mutations. Nat Genet. 2018;50:1219–24.
116. Zhang JP, Robinson D, Yu J, Gallego J, Fleischhacker WW, Kahn RS, et al. Schi- 142. Martin AR, Kanai M, Kamatani Y, Okada Y, Neale BM, Daly MJ. Clinical use of current
zophrenia polygenic risk score as a predictor of antipsychotic efficacy in first- polygenic risk scores may exacerbate health disparities. Nat Genet. 2019;51:584–91.
episode psychosis. Am J Psychiatry. 2019;176:21–8. 143. Curtis D. Polygenic risk score for schizophrenia is more strongly associated with
117. Landi I, Kaji DA, Cotter L, Van Vleck T, Belbin G, Preuss M, et al. Prognostic value ancestry than with schizophrenia. Psychiatr Genet. 2018;28:85–9.
of polygenic risk scores for adults with psychosis. Nat Med. 2021;27:1576–81. 144. World Health Organization. Global burden of mental disorders and the need for
118. Phan BN, Bohlen JF, Davis BA, Ye Z, Chen HY, Mayfield B, et al. A myelin-related a comprehensive, coordinated response from health and social workers at the
transcriptomic profile is shared by Pitt-Hopkins syndrome models and human country level: report by the Secretariat. 2012.
autism spectrum disorder. Nat Neurosci. 2020;23:375–85.
119. Zerbi V, Pagani M, Markicevic M, Matteoli M, Pozzi D, Fagiolini M, et al. Brain
mapping across 16 autism mouse models reveals a spectrum of functional
connectivity subtypes. Mol Psychiatry. 2021;26:7610–20.
ACKNOWLEDGEMENTS
120. Kelly E, Meng F, Fujita H, Morgado F, Kazemi Y, Rice LC, et al. Regulation of
Parts of Fig. 1 were drawn by using pictures from Servier Medical Art. Servier Medical
autism-relevant behaviors by cerebellar-prefrontal cortical circuits. Nat Neurosci.
Art by Servier is licensed under a Creative Commons Attribution 3.0 Unported License
2020;23:1102–10.
(https://creativecommons.org/licenses/by/3.0/). This work was supported by grants
121. Paulsen B, Velasco S, Kedaigle AJ, Pigoni M, Quadrato G, Deo AJ, et al. Autism
from the Japan Agency for Medical Research and Development (AMED) under Grant
genes converge on asynchronous development of shared neuron classes.
Number JP21km0405214 (to AT) and JSPS KAKENHI under Grant Numbers
Nature. 2022;602:268–73.
JP20H05777 (to AT), 21H02855 (to AT), 22H02991 (to AT), and 21K15752 (to TN).
122. Fromer M, Roussos P, Sieberts SK, Johnson JS, Kavanagh DH, Perumal TM, et al.
Gene expression elucidates functional impact of polygenic risk for schizo-
phrenia. Nat Neurosci. 2016;19:1442–53.
123. Marek S, Tervo-Clemmens B, Calabro FJ, Montez DF, Kay BP, Hatoum AS, et al.
Reproducible brain-wide association studies require thousands of individuals. AUTHOR CONTRIBUTIONS
Nature. 2022;603:654–60. TN and AT performed investigation and wrote the manuscript.
22
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The molecular pathology of schizophrenia: an overview of

© The Author(s) 2023

Molecular Psychiatry; https://doi.org/10.1038/s41380-023-02005-2

INTRODUCTION the critical problem of multiple testing and relevant statistical

Received: 1 August 2022 Revised: 15 February 2023 Accepted: 15 February 2023

PDZK1P2, PDZK1P2, NBPF8, NBPF8, FAM108A3, AL049742.8-2, NBPF12, PRKAB2,

TFRC, ZDHHC19, AC069257.28-2, PCYT1A, TM4SF19, TCTEX1D2, UBXN7, C3orf43,

ATXN2L, TUFM, SH2B1, ATP2A1, RABEP2, CD19, NFATC2IP, SPNS1, LAT,

WBSCR1, LAT2, RFC2, CYLN2, GTF2IRD1, GTF2I

of rare coding variants in other neurodevelopmental disorders

MFI2, DLG1, BDH1

through large-scale studies, which underlie the genetic pathology

of schizophrenia, in Table 1, together with the landscape of their

studies. To more accurately estimate their effect sizes and the

robustness of the association, further larger studies are always

Functional convergence of the ﬁndings in genetic studies

As described above, genetic studies have identiﬁed a number of

statistically robust new genes and variants; however, these in

further analysis is needed to translate genetic ﬁndings into the

by utilizing MAGMA [34] and DNENRICH [16] that appropriately

disequilibrium. We, therefore, examined how the results of these

analysis for 7315 and 1491 terms are available, respectively. Of

showed signiﬁcant enrichment at uncorrected P < 0.05 in both of

these two studies (Fig. 2a). The statistical signiﬁcance (−log10 P

**Odds ratio for LOF/PTV and MPC ≥ 3 missense variants.

correlation coefﬁcient = 0.39, P = 6.67 × 10−54). This result indi-

cates that there is a convergence of molecular and biological

pathways implicated by common SNPs and rare deleterious

channels and synaptic transmissions, were signiﬁcant after

Bonferroni correction in both the PGC GWAS and the SCHEMA

above) were visualized as networks by connecting them based

transporter activities; neuronal components (synapse, axon, and

dendrite); chromatin or histone organization. Though the cluster

of chromatin or histone organization was only supported by

SCHEMA, the former two clusters (“channel or transporter

enrichment in both studies. Taken together, these can be

considered molecular pathways whose involvement in

the frontal synchrony in

Heterozygous B6129SF1/J 65 Yin X. 2021 • Delayed spine Delayed

duplication of chr7 signaling cued increased

• Enrichment of PFC and striatal

b+/-) NRXN1α/β+/- epigenetic regulation multi-electrode in-frame human

2p16.3 (NRXN1) deletion with OR smaller than ten [30, 32].

considered as speciﬁcally modeling schizophrenia. Third, particu-

larly in the case of CNV-based animal models, evaluation of the

between rodents and humans. Lastly, we would like to emphasize

that a number of valuable ﬁndings interpreted as being relevant

to the molecular pathology of schizophrenia have also been

obtained from analyses of models in which genes not strongly

supported by currently available evidence from human genetics

dysregulation of the monoaminergic system is involved in the

pathophysiology of schizophrenia [108, 109], and therefore

form a foundation not only for the study of schizophrenia patients

not only those in genome-wide signiﬁcant loci, it is clear that there

the stringent signiﬁcance threshold with the current sample sizes.

using animal and cellular models with modiﬁcations of such

homozygous mutants, even though heterozygous variants are