TNNT1

Template:PBB The TNNT1 gene is located at 19q13.4 in the human chromosomal genome, encoding the slow twitch skeletal muscle isoform of troponin T (ssTnT). ssTnT is an ~32-kDa protein consisting of 262 amino acids (including the first methionine) with an isoelectric point (pI) of 5.95. It is the tropomyosin binding and thin filament anchoring subunit of the troponin complex in the sarcomeres of slow twitch skeletal muscle fibers (Perry 1998; Jin, Zhang et al. 2008; Wei and Jin 2011). TNNT1 gene is specifically expressed in slow skeletal muscle of vertebrates, with one exception that dry land toad (Bufo) cardiac muscle expresses ssTnT other than  cardiac TnT (Feng, Chen et al. 2012).

Evolution of TnT isoform genes

Three homologous genes have evolved in vertebrates, encoding three muscle type specific isoforms of TnT (Wei and Jin 2011). Each of the TnT isoform genes is linked to one of the three troponin I (Dweck, Sanchez-Gonzalez et al.) isoform genes encoding the inhibitory subunit of the troponin complex, in chromosomal DNA to form three gene pairs: The fast skeletal muscle TnI (fsTnI)-fsTnT, ssTnI-cardiac (cTnT) and cTnI-ssTnT gene pairs. Sequence and epitope conservation studies suggested that genes encoding the muscle type specific TnT and TnI isoforms may have evolved from duplications of a fsTnI-like-fsTnT-like gene pair (Chong and Jin 2009). Evolutionary lineage of the three TnI-TnT gene pairs shows that cTnI-ssTnT is the newest (Chong and Jin 2009) and most closely linked (Huang and Jin 1999) pair (Fig. 1). 

Protein sequence alignment demonstrated that TNNT1 genes are highly conserved among vertebrate species (Fig. 2), especially in the middle and C-terminal regions, while the three muscle type isoforms are significantly diverged among vertebrate species (Jin, Zhang et al. 2008; Wei and Jin 2011).  

Alternative splicing

In mammalian and avian species, TNNT1 gene has a total of 14 exons, among which exon 5 encoding an 11-amino acid in the N-terminal region is alternatively spliced, generating a high molecular weight and a low molecular weight slow TnT splice forms (Gahlmann, Troutt et al. 1987; Jin, Chen et al. 1998; Huang, Chen et al. 1999). Biochemical studies showed that TnT splice forms have detectable different molecular conformation in the middle and C-terminal regions, producing different binding affinities for TnI and tropomyosin (Jin, Zhang et al. 2008; Wei and Jin 2011). The alternative splice forms of ssTnT play a role in skeletal muscle adaptation in physiologic and pathological conditions (Larsson, Wang et al. 2008). Alternative splicing at alternative acceptor sites of intron 5 generates a single amino acid difference in the N-terminal region of ssTnT (Huang, Chen et al. 1999), of which functional significance has not been established. 

Clinical significance

A nonsense mutation E180X in the exon 11 of TNNT1 gene causes Amish Nemaline Myopathy (ANM), which is a severe form of recessive nemaline myopathy originally found in the Old Order Amish population in Pennsylvania, USA (Johnston, Kelley et al. 2000; Jin, Brotto et al. 2003). Truncation of the ssTnT polypeptide chain by the E180X mutation deletes the tropomyosin-binding site 2 (Jin and Chong 2010) as well as the binding sites for TnI and troponin C (TnC) in the C-terminal region (Fig. 3). Consistent with the recessive phenotype, the truncated ssTnT is incapable of incorporation into the myofilaments and completely degraded in muscle cells (Martin 1981) .  Tnnt1 gene targeted mouse studies reproduced the myopathic phenotypes of ANM (Feng, Wei et al. 2009; Wei, Lu et al. 2014). ssTnT null mice showed significantly decreased type I slow fibers in diaphragm and soleus muscles with hypertrophy of type II fast fibers, increased fatigability, and active regeneration of slow fibers (Feng, Wei et al. 2009; Wei, Lu et al. 2014). 

Recent case reports identified three more mutations in TNNT1 gene to cause nemaline myopathies outside the Amish population. A nonsense mutation S108X in exon 9 was identified in a Hispanic male patient with severe recessive nemaline myopathy phenotype (Marra, Engelstad et al. 2015). A Dutch patient with compound heterozygous TNNT1 gene mutations that cause exon 8 and exon 14 deletions also presents nemaline myopathy phenotypes (van der Pol, Leijenaar et al. 2014). A rearrangement in TNNT1 gene (c.574_577 delins TAGTGCTGT) leading to aberrant splicing that causes C-terminal truncation of the protein (L203 truncation) was reported in 9 Palestinian patients from 7 unrelated families with recessively inherited nemaline Myopathy (Abdulhaq, Daana et al. 2015).

Illustrated in Fig. 3, the S108X mutation truncates ssTnT protein to cause a loss of functional structures equivalent to that of E180X. The exon 8 deletion destructs the middle region tropomyosin-binding site 1 (Jin and Chong 2010; Amarasinghe and Jin 2015). The L203 truncation deletes the binding sites for TnI and TnC but preserves both tropomyosin-binding sites 1 and 2 (Jin and Chong, 2010) It remains to be invistigated whether this novel mutation is able to bind the actin-tropomyosin thin filament in vivo and how it causes recessive nemaline myopathy.

Alternative splicing of exon 5 produces high and low molecular weight splice forms of ssTnT. The low molecular ssTnT was significantly upregulated in type 1 (demyelination) but not type 2 (axon loss) Charcot-Marie-Tooth disease (Larsson, Wang et al. 2008), suggesting that structural modification of TnT in the myofilaments may contribute to adaptation to abnormalities in neuronal activation. 

References

Abdulhaq, U. N., M. Daana, et al. (2015). "Nemaline body myopathy caused by a novel mutation in Troponin T1 (TNNT1)." Muscle Nerve.

Amarasinghe, C. and J. P. Jin (2015). "N-Terminal Hypervariable Region of Muscle Type Isoforms of Troponin T Differentially Modulates the Affinity of Tropomyosin-Binding Site 1." Biochemistry 54(24): 3822-3830.

Chong, S. M. and J. P. Jin (2009). "To investigate protein evolution by detecting suppressed epitope structures." J Mol Evol 68(5): 448-460.

Dweck, D., M. A. Sanchez-Gonzalez, et al. (2014). "Long term ablation of protein kinase A (PKA)-mediated cardiac troponin I phosphorylation leads to excitation-contraction uncoupling and diastolic dysfunction in a knock-in mouse model of hypertrophic cardiomyopathy." J Biol Chem 289(33): 23097-23111.

Feng, H. Z., X. Chen, et al. (2012). "Toad heart utilizes exclusively slow skeletal muscle troponin T: an evolutionary adaptation with potential functional benefits." J Biol Chem 287(35): 29753-29764.

Feng, H. Z., B. Wei, et al. (2009). "Deletion of a genomic segment containing the cardiac troponin I gene knocks down expression of the slow troponin T gene and impairs fatigue tolerance of diaphragm muscle." J Biol Chem 284(46): 31798-31806.

Gahlmann, R., A. B. Troutt, et al. (1987). "Alternative splicing generates variants in important functional domains of human slow skeletal troponin T." J Biol Chem 262(33): 16122-16126.

Huang, Q. Q., A. Chen, et al. (1999). "Genomic sequence and structural organization of mouse slow skeletal muscle troponin T gene." Gene 229(1-2): 1-10.

Huang, Q. Q. and J. P. Jin (1999). "Preserved close linkage between the genes encoding troponin I and troponin T, reflecting an evolution of adapter proteins coupling the Ca(2+) signaling of contractility." J Mol Evol 49(6): 780-788.

Jin, J. P., M. A. Brotto, et al. (2003). "Truncation by Glu180 nonsense mutation results in complete loss of slow skeletal muscle troponin T in a lethal nemaline myopathy." J Biol Chem 278(28): 26159-26165.

Jin, J. P., A. Chen, et al. (1998). "Three alternatively spliced mouse slow skeletal muscle troponin T isoforms: conserved primary structure and regulated expression during postnatal development." Gene 214(1-2): 121-129.

Jin, J. P. and S. M. Chong (2010). "Localization of the two tropomyosin-binding sites of troponin T." Arch Biochem Biophys 500(2): 144-150.

Jin, J. P., Z. Zhang, et al. (2008). "Isoform diversity, regulation, and functional adaptation of troponin and calponin." Crit Rev Eukaryot Gene Expr 18(2): 93-124.

Johnston, J. J., R. I. Kelley, et al. (2000). "A novel nemaline myopathy in the Amish caused by a mutation in troponin T1." Am J Hum Genet 67(4): 814-821.

Larsson, L., X. Wang, et al. (2008). "Adaptation by alternative RNA splicing of slow troponin T isoforms in type 1 but not type 2 Charcot-Marie-Tooth disease." Am J Physiol Cell Physiol 295(3): C722-731.

Marra, J. D., K. E. Engelstad, et al. (2015). "Identification of a novel nemaline myopathy-causing mutation in the troponin T1 (TNNT1) gene: a case outside of the old order Amish." Muscle Nerve 51(5): 767-772.

Martin, A. F. (1981). "Turnover of cardiac troponin subunits. Kinetic evidence for a precursor pool of troponin-I." J Biol Chem 256(2): 964-968.

Palm, T., S. Graboski, et al. (2001). "Disease-causing mutations in cardiac troponin T: identification of a critical tropomyosin-binding region." Biophys J 81(5): 2827-2837.

Perry, S. V. (1998). "Troponin T: genetics, properties and function." J Muscle Res Cell Motil 19(6): 575-602.

van der Pol, W. L., J. F. Leijenaar, et al. (2014). "Nemaline myopathy caused byTNNT1 mutations in a Dutch pedigree." Mol Genet Genomic Med 2(2): 134-137.

Vinogradova, M. V., D. B. Stone, et al. (2005). "Ca(2+)-regulated structural changes in troponin." Proc Natl Acad Sci U S A 102(14): 5038-5043.

Wei, B. and J. P. Jin (2011). "Troponin T isoforms and posttranscriptional modifications: Evolution, regulation and function." Arch Biochem Biophys 505(2): 144-154.

Wei, B., Y. Lu, et al. (2014). "Deficiency of slow skeletal muscle troponin T causes atrophy of type I slow fibres and decreases tolerance to fatigue." J Physiol 592(Pt 6): 1367-1380.

External links

• GeneReviews/NCBI/NIH/UW entry on Nemaline Myopathy