Search Results (30)

Calmodulins (CaMs) and calmodulin-like proteins (CMLs) belong to families of calcium-sensors that act as calcium ion (Ca²⁺) signal-decoding proteins and regulate downstream target proteins. As a tropical halophyte, Canavalia rosea shows great resistance to multiple abiotic stresses, including high salinity/alkalinity, extreme drought, heat, and intense sunlight. However, investigations of calcium ion signal transduction involved in the stress responses of C. rosea are limited. The CaM and CML gene families have been identified and characterized in many other plant species. Nevertheless, there is limited available information about these genes in C. rosea. In this study, a bioinformatic analysis, including the gene structures, conserved protein domains, phylogenetic relationships, chromosome distribution, and gene synteny, was comprehensively performed to identify and characterize CrCaMs and CrCMLs. A spatio-temporal expression assay in different organs and environmental conditions was then conducted using the RNA sequencing technique. Additionally, several CrCaM and CrCML members were then cloned and functionally characterized using the yeast heterogeneous expression system, and some of them were found to change the tolerance of yeast to heat, salt, alkalinity, and high osmotic stresses. The results of this study provide a foundation for understanding the possible roles of the CrCaM and CrCML genes, especially for halophyte C. rosea’s natural ecological adaptability for its native habitats. This study also provides a theoretical basis for further study of the physiological and biochemical functions of plant CaMs and CMLs that are involved in tolerance to multiple abiotic stresses. Full article

Cuttage is the main propagation method of tea plant cultivars in China. However, some tea softwood cuttings just form an expanded and loose callus at the base, without adventitious root (AR) formation during the propagation period. Meanwhile, exogenous auxin could promote the AR formation of tea plant cuttings, but the regulation mechanism has not yet explained clearly. We conducted this study to elucidate the regulatory mechanism of exogenous auxin-induced adventitious root (AR) formation of such cuttings. The transcriptional expression profile of non-rooting tea calluses in response to exogenous IBA and NAA was analyzed using ONT RNA Seq technology. In total, 56,178 differentially expressed genes (DEGs) were detected, and most of genes were significantly differentially expressed after 12 h of exogenous auxin treatment. Among these DEGs, we further identified 80 DEGs involved in the auxin induction pathway and AR formation. Specifically, 14 auxin respective genes (ARFs, GH3s, and AUX/IAAs), 3 auxin transporters (AUX22), 19 auxin synthesis- and homeostasis-related genes (cytochrome P450 (CYP450) and calmodulin-like protein (CML) genes), and 44 transcription factors (LOB domain-containing protein (LBDs), SCARECROW-LIKE (SCL), zinc finger protein, WRKY, MYB, and NAC) were identified from these DEGs. Moreover, we found most of these DEGs were highly up-regulated at some stage before AR formation, suggesting that they may play a potential role in the AR formation of tea plant cuttings. In summary, this study will provide a theoretical foundation to deepen our understanding of the molecular mechanism of AR formation in tea cuttings induced by auxin during propagation time. Full article

Soybean mosaic virus (SMV) is one of the main pathogens that can negatively affect soybean production and quality. To study the gene regulatory network of soybeans in response to SMV SC15, the resistant line X149 and susceptible line X97 were subjected to transcriptome analysis at 0, 2, 8, 12, 24, and 48 h post-inoculation (hpi). Differential expression analysis revealed that 10,190 differentially expressed genes (DEGs) responded to SC15 infection. Weighted gene co-expression network analysis (WGCNA) was performed to identify highly related resistance gene modules; in total, eight modules, including 2256 DEGs, were identified. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of 2256 DEGs revealed that the genes significantly clustered into resistance-related pathways, such as the plant–pathogen interaction pathway, mitogen-activated protein kinases (MAPK) signaling pathway, and plant hormone signal transduction pathway. Among these pathways, we found that the flg22, Ca²⁺, hydrogen peroxide (H₂O₂), and abscisic acid (ABA) regulatory pathways were fully covered by 36 DEGs. Among the 36 DEGs, the gene Glyma.01G225100 (protein phosphatase 2C, PP2C) in the ABA regulatory pathway, the gene Glyma.16G031900 (WRKY transcription factor 22, WRKY22) in Ca²⁺ and H₂O₂ regulatory pathways, and the gene Glyma.04G175300 (calcium-dependent protein kinase, CDPK) in Ca²⁺ regulatory pathways were highly connected hub genes. These results indicate that the resistance of X149 to SC15 may depend on the positive regulation of flg22, Ca²⁺, H₂O₂, and ABA regulatory pathways. Our study further showed that superoxide dismutase (SOD) activity, H₂O₂ content, and catalase (CAT) and peroxidase (POD) activities were significantly up-regulated in the resistant line X149 compared with those in 0 hpi. This finding indicates that the H₂O₂ regulatory pathway might be dependent on flg22- and Ca²⁺-pathway-induced ROS generation. In addition, two hub genes, Glyma.07G190100 (encoding F-box protein) and Glyma.12G185400 (encoding calmodulin-like proteins, CMLs), were also identified and they could positively regulate X149 resistance. This study provides pathways for further investigation of SMV resistance mechanisms in soybean. Full article

Calmodulin (CaM) and calmodulin-like (CML) proteins are major Ca²⁺ sensors involved in the regulation of plant development and stress responses by converting Ca²⁺ signals into appropriate cellular responses. However, characterization and expression analyses of CaM/CML genes in the precious species, Phoebe bournei, remain limited. In this study, five PbCaM and sixty PbCML genes were identified that only had EF-hand motifs with no other functional domains. The phylogenetic tree was clustered into 11 subgroups, including a unique clade of PbCaMs. The PbCaMs were intron-rich with four EF-hand motifs, whereas PbCMLs had two to four EF-hands and were mostly intronless. PbCaMs/CMLs were unevenly distributed across the 12 chromosomes of P. bournei and underwent purifying selection. Fragment duplication was the main driving force for the evolution of the PbCaM/CML gene family. Cis-acting element analysis indicated that PbCaMs/CMLs might be related to hormones, growth and development, and stress response. Expression analysis showed that PbCaMs were generally highly expressed in five different tissues and under drought stress, whereas PbCMLs showed specific expression patterns. The expression levels of 11 candidate PbCaMs/CMLs were responsive to ABA and MeJA, suggesting that these genes might act through multiple signaling pathways. The overexpression of PbCaM3/CML13 genes significantly increased the tolerance of yeast cells to drought stress. The identification and characterization of the CaM/CML gene family in P. bournei laid the foundation for future functional studies of these genes. Full article

Calmodulin-like proteins (CMLs) are an important family of plant calcium sensor proteins that sense and decode changes in the intracellular calcium concentration in response to environmental and developmental stimuli. Nonetheless, the specific functions of individual CML family members remain largely unknown. This study aims to explore the role of the Vitis amurensis VaCML92 gene in the development of its high stress resistance and the production of stilbenes. The expression of VaCML92 was sharply induced in V. amurensis cuttings after cold stress. The VaCML92 gene was cloned and its role in the abiotic stress responses and stilbene production in grapevine was further investigated. The VaCML92-overexpressing callus cell cultures of V. amurensis and soil-grown plants of Arabidopsis thaliana exhibited enhanced tolerance to cold stress and, to a lesser extent, to the drought, while their tolerance to heat stress and high salinity was not affected. In addition, the overexpression of VaCML92 increased stilbene production in the V. amurensis cell cultures by 7.8–8.7-fold. Taken together, the data indicate that the VaCML92 gene is involved as a strong positive regulator in the rapid response to cold stress, the induction of cold stress resistance and in stilbene production in wild grapevine. Full article

As significant Ca²⁺ sensors, calmodulin (CaM) and calmodulin-like proteins (CML), have been associated with a variety of environmental conditions in plants. However, whether CaMs/CMLs are related to the stress of phytoplasma infection has not been reported in Paulownia fortunei. In the current study, 5 PfCaMs and 58 PfCMLs were detected through a genome-wide investigation. The number of EF-hand motifs in all PfCaMs/CMLs varied. Bioinformatics analyses, including protein characteristics, conserved domain, gene structure, cis-elements, evolutionary relationship, collinearity, chromosomal location, post-translation modification site, subcellular localization and expression pattern analyses, represented the conservation and divergence of PfCaMs/CMLs. Furthermore, some PfCaMs/CMLs might be involved in plants’ reaction to phytoplasma infection and exogenous calcium therapy, indicating these genes may play a role in abiotic as well as biotic stress responses. In addition, subcellular localization analysis showed that PfCML10 was located in the cell membrane and nucleus. In summary, these findings establish a stronger platform for their subsequent functional investigation in trees and further characterize their roles in Paulownia witches’ broom (PaWB) occurrence. Full article

The jujube witches’ broom (JWB) disease is a severe threat to jujube trees, with only a few cultivars being genuinely tolerant or resistant to phytoplasma. The defense mechanism of jujube trees against phytoplasma is still unclear. In this study, we aimed to investigate the tolerance mechanism of Indian jujube ‘Cuimi’ to JWB and identify the key genes that contribute to JWB high tolerance. Based on the symptoms and phytoplasma concentrations after infection, we confirmed the high tolerance of ‘Cuimi’ to JWB. Comparative transcriptome analysis was subsequently performed between ‘Cuimi’ and ‘Huping’, a susceptible cultivar of Chinese jujube. Unique gene ontology (GO) terms were identified in ‘Cuimi’, such as protein ubiquitination, cell wall biogenesis, cell surface receptor signaling pathway, oxylipin biosynthetic process, and transcription factor activity. These terms may relate to the normal development and growth of ‘Cuimi’ under phytoplasma infection. We identified 194 differential expressed genes related to JWB high tolerance, involved in various processes, such as reactive oxygen species (ROS), Ca²⁺ sensors, protein kinases, transcription factors (TFs), lignin, and hormones. Calmodulin-like (CML) genes were significantly down-regulated in infected ‘Cuimi’. We speculated that the CML gene may act as a negative regulatory factor related to JWB high tolerance. Additionally, the cinnamoyl-CoA reductase-like SNL6 gene was significantly up-regulated in infected ‘Cuimi’, which may cause lignin deposition, limit the growth of phytoplasma, and mediate immune response of ‘Cuimi’ to phytoplasma. Overall, this study provides insights into the contribution of key genes to the high tolerance of JWB in Indian jujube ‘Cuimi’. Full article

Calmodulins (CaMs) and Calmodulin-like proteins (CMLs) are vital in plant growth, development, and stress responses. However, CaMs and CMLs have not been fully identified and characterized in brown algae, which has been evolving independently of the well-studied green plant lineage. In this study, whole-genome searches revealed one SjCaM and eight SjCMLs in Saccharina japonica, and one EsCaM and eleven EsCMLs in Ectocarpus sp. SjCaM and EsCaM encoded identical protein products and shared 88.59–89.93% amino acid identities with Arabidopsis thaliana AtCaMs, thereby indicating that brown algae CaMs retained a similar Ca²⁺ sensors function as in plants. The phylogenetic and gene structure analysis results showed that there was significant divergence in the gene sequences among brown algae CMLs. Furthermore, evolutionary analysis indicated that the function of brown alga CMLs was relatively conserved, which may be related to the fact that brown algae do not need to face complex environments like terrestrial plants. Regulatory elements prediction and the expression analysis revealed the probable functioning of SjCaM/CML genes in gametophyte development and the stress response in S. japonica. In addition, the SjCaM/SjCMLs interacting proteins and chemicals were preliminarily predicted, suggesting that SjCaM/SjCMLs might play putative roles in Ca²⁺/CaM-mediated growth and development processes and stimulus responses. Therefore, these results will facilitate our understanding of the evolution of brown algae CaMs/CMLs and the functional identification of SjCaM/SjCMLs. Full article

MicroRNAs (miRNAs) belong to non-coding small RNAs which have been shown to take a regulatory function at the posttranscriptional level in plant growth development and response to abiotic stress. Hemerocallis fulva is an herbaceous perennial plant with fleshy roots, wide distribution, and strong adaptability. However, salt stress is one of the most serious abiotic stresses to limit the growth and production of Hemerocallis fulva. To identify the miRNAs and their targets involved in the salt stress resistance, the salt-tolerant H. fulva with and without NaCl treatment were used as materials, and the expression differences of miRNAs–mRNAs related to salt-tolerance were explored and the cleavage sites between miRNAs and targets were also identified by using degradome sequencing technology. In this study, twenty and three significantly differential expression miRNAs (p-value < 0.05) were identified in the roots and leaves of H. fulva separately. Additionally, 12,691 and 1538 differentially expressed genes (DEGs) were also obtained, respectively, in roots and leaves. Moreover, 222 target genes of 61 family miRNAs were validated by degradome sequencing. Among the DE miRNAs, 29 pairs of miRNA targets displayed negatively correlated expression profiles. The qRT-PCR results also showed that the trends of miRNA and DEG expression were consistent with those of RNA-seq. A gene ontology (GO) enrichment analysis of these targets revealed that the calcium ion pathway, oxidative defense response, microtubule cytoskeleton organization, and DNA binding transcription factor responded to NaCl stress. Five miRNAs, miR156, miR160, miR393, miR166, and miR396, and several hub genes, squamosa promoter-binding-like protein (SPL), auxin response factor 12 (ARF), transport inhibitor response 1-like protein (TIR1), calmodulin-like proteins (CML), and growth-regulating factor 4 (GRF4), might play central roles in the regulation of NaCl-responsive genes. These results indicate that non-coding small RNAs and their target genes that are related to phytohormone signaling, Ca²⁺ signaling, and oxidative defense signaling pathways are involved in H. fulva’s response to NaCl stress. Full article

The calmodulin-like (CML) family is an important calcium (Ca²⁺) sensor in plants and plays a pivotal role in the response to abiotic and biotic stresses. As one of the most salt-tolerant grass species, Paspalums vaginatum is resistant to multiple abiotic stresses, such as salt, cold, and drought. However, investigations of PvCML proteins in P. vaginatum have been limited. Based on the recently published P. vaginatum genome, we identified forty-nine PvCMLs and performed a comprehensive bioinformatics analysis of PvCMLs. The main results showed that the PvCMLs were unevenly distributed on all chromosomes and that the expansion of PvCMLs was shaped by tandem and segmental duplications. In addition, cis-acting element analysis, expression profiles, and qRT–PCR analysis revealed that PvCMLs were involved in the response to salt and cold stress. Most interestingly, we found evidence of a tandem gene cluster that independently evolved in P. vaginatum and may participate in cold resistance. In summary, our work provides important insight into how grass species are resistant to abiotic stresses such as salt and cold and could be the basis of further gene function research on CMLs in P. vaginatum. Full article

Sclerotinia stem rot caused by Sclerotinia sclerotiorum is one of the most destructive diseases in Brassica rapa. Verticillium dahliae Aspf2-like protein (VDAL) is a secretory protein of V. dahliae which has been shown to enhance the resistance against fungal infections in several plants. Nonetheless, the molecular mechanisms of VDAL-primed disease resistance are still poorly understood. In this study, we performed physiological, biochemical, and transcriptomic analyses of Brassica rapa in order to understand how VDAL confers resistance to S. sclerotiorumn infections in plants. The results showed that foliar application of VDAL significantly reduced the plaque area on leaves inoculated with S. sclerotiorum. It also enhanced antioxidant capacity by increasing activities of superoxide dismutase (SOD), peroxidase (POD), peroxidase (APX), glutathione reductase (GR), protoporphyrinogen oxidase (PPO), and defense-related enzymes β-1,3-glucanase and chitinase during the infection periods. This occurred in parallel with significantly reduced relative conductivity at different periods and lower malondialdehyde (MDA) content as compared to sole S. sclerotiorum inoculation. Transcriptomic analysis showed a total of 146 (81 up-regulated and 65 down-regulated) differentially expressed genes (DEGs) in VDAL-treated leaves compared to the control. The most enriched three Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were the mitogen–activated protein kinase (MAPK) signaling pathway, plant hormone signal transduction, and plant-pathogen interaction, all of which were associated with plant immunity. DEGs associated with MAPK and hormone signal transduction pathways were ethylene response sensor ERS2, EIN3 (Ethylene Insensitive3)-binding F-box protein 2 (EBF2), ethylene-responsive transcription factor ERF94, MAPK 9 (MKK9), protein phosphatase 2C (PP2C37), auxin-responsive proteins (AUX/IAA1 and 19), serine/threonine-protein kinase CTR1, and abscisic acid receptors (PLY 4 and 5). Among the DEGs linked with the plant–pathogen interaction pathway were calmodulin-like proteins (CML5, 24, 27), PTI1-like tyrosine protein kinase 3 (Pti13) and transcription factor MYB30, all of which are known to play key roles in pathogen-associated molecular pattern (PAMP)-triggered immunity and effector-triggered immunity (ETI) for hypersensitive response (HR), cell wall reinforcement, and stomatal closure in plants. Overall, VDLA treatment triggered repression of the auxin and ABA signaling pathways and de-repression of the ethylene signaling pathways in young B. rapa seedlings to increase plant innate immunity. Our results showed that VDAL holds great potential to enhance fungal disease resistance in B. rapa crop. Full article

As an important Ca²⁺ sensor, calmodulin (CaM) and calmodulin-like protein (CML) play core roles in plant growth, development, and response to environmental stimuli. The CaM/CML gene family has been well characterized in various plant species, such as Arabidopsis thaliana, rice, and tomato; however, in the tea plant, the CaM/CML gene family has not been systematically and comprehensively characterized. In the present study, a total of 5 CsCaM and 60 CsCML proteins were identified from the tea plant genome, which were unevenly distributed on the 14 chromosomes of the tea plant. All the proteins contained two to four EF-hand domains. Meanwhile, an integrated analysis of physicochemical properties, sequence structure, motif identification, phylogeny, gene duplication, promoter cis-elements, and RNA-seq expression profiles in the CsCaM/CML gene family was performed. Transcriptome analysis revealed that CsCaM/CMLs were differentially expressed in different tissues of the tea plant, suggesting their potential roles in plant growth and development. The expression profiles associated with various stress treatments revealed that CsCaM/CML genes were involved in a wide range of abiotic factors, including cold and drought stress. Quantitative real-time PCR (qRT-PCR) was also used to validate the differences in expression under abiotic stress. Overall, these findings enhanced our understanding of CsCaM/CML genes and provided useful information for further research into their molecular functions in abiotic stress response, and in multiple physiological processes in the tea plant. Full article

Calmodulin (CaM) and calmodulin-like proteins (CML) act as significant Ca²⁺ sensors binding Ca²⁺ with EF-hand motifs and have been reported to be involved in various environmental stresses in plants. In this study, calmodulin CsCaM/CML gene family members were identified based on the genome of Chrysanthemum seticuspe published recently; a phylogenetic tree was constructed; gene structures and chromosomal locations of CsCaM/CML were depicted; cis-acting regulatory elements were predicted; collinearity and duplicate events of CaM/CML were analyzed using MCScanX software; and the expression levels of CsCaM/CML in response to abiotic stress were analyzed, based on the published RNA-seq data. We identified 86 CsCaM/CML (4 CsCaMs and 82 CsCMLs) genes in total. Promoter sequences of CsCaM/CML contained elements related to abiotic stresses (including low-temperature and anaerobic stresses) and plant hormones (including abscisic acid (ABA), MeJA, and salicylic acid). CsCaM/CML genes were distributed on nine chromosomes unevenly. Collinearity analysis indicated that recent segmental duplications significantly enlarged the scale of the CML family in C. seticuspe. Four CsCMLs (CsCML14, CsCML50, CsCML65, and CsCML79) were statistically differentially regulated under low-temperature and salt stress compared with those in the normal condition. These results indicate diverse roles of CsCaM/CML in plant development and in response to environmental stimuli in C. seticuspe. Full article

Calmodulins (CAMs) and calmodulin-like proteins (CMLs) can participate in the regulation of various physiological processes via sensing and decoding Ca²⁺ signals. To reveal the characteristics of the CAM/CML family in Ginkgo biloba, a comprehensive analysis was performed at the genome-wide level. A total of 26 CAMs/CMLs, consisting of 5 GbCAMs and 21 GbCMLs, was identified on 11 out of 12 chromosomes in G. biloba. They displayed a certain degree of multiplicity in their sequences, albeit with conserved EF hands. Collinearity analysis suggested that tandem rather than segmental or whole-genome duplications were likely to play roles in the evolution of the Ginkgo CAM/CML family. Furthermore, GbCAMs/GbCMLs were grouped into higher, lower, and moderate expression in magnitude. The cis-acting regulatory elements involved in phytohormone-responsiveness within GbCAM/GbCML promotors may explain their varied expression profiles. The ectopic expression of a GbCML gene (Gb_30819) in transgenic Arabidopsis led to phenotypes with significantly shortened root length and seedling height, and decreased yields of both pods and seeds. Moreover, an electrophoresis mobility shift assay demonstrated the Ca²⁺-binding activity of Gb_30819 in vitro. Altogether, these results contribute to insights into the characteristics of the evolution and expression of GbCAMs/GbCMLs, as well as evidence for Ca²⁺-CAM/CML pathways functioning within the ancient gymnosperm G. biloba. Full article

Stilbenes are plant phenolics known to rapidly accumulate in grapevine and other plants in response to injury or pathogen attack and to exhibit a great variety of healing beneficial effects. It has previously been shown that several calmodulin-like protein (CML) genes were highly up-regulated in cell cultures of wild-growing grapevine Vitis amurensis Rupr. in response to stilbene-modulating conditions, such as stress hormones, UV-C, and stilbene precursors. Both CML functions and stilbene biosynthesis regulation are still poorly understood. In this study, we investigated the effect of overexpression of five VaCML genes on stilbene and biomass accumulation in the transformed cell cultures of V. amurensis. We obtained 16 transgenic cell lines transformed with the VaCML52, VaCML65, VaCML86, VaCML93, and VaCML95 genes (3–4 independent lines per gene) under the control of the double CaMV 35S promoter. HPLC-MS analysis showed that overexpression of the VaCML65 led to a considerable and consistent increase in the content of stilbenes of 3.8–23.7 times in all transformed lines in comparison with the control calli, while biomass accumulation was not affected. Transformation of the V. amurensis cells with other analyzed VaCML genes did not lead to a consistent and considerable effect on stilbene biosynthesis in the cell lines. The results indicate that the VaCML65 gene is implicated in the signaling pathway regulating stilbene biosynthesis as a strong positive regulator and can be useful in viticulture and winemaking for obtaining grape cultivars with a high content of stilbenes and stress resistance. Full article