Slow skeletal muscle troponin T (sTnT) is a protein that in humans is encoded by the TNNT1gene.[5][6]
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.[7][8][9] 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.[10]
Evolution
Three homologous genes have evolved in vertebrates, encoding three muscle type specific isoforms of TnT.[9] Each of the TnT isoform genes is linked to one of the three troponin I[11] 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.[12] Evolutionary lineage of the three TnI-TnT gene pairs shows that cTnI-ssTnT is the newest[12] and most closely linked.[13]
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.[8][9]
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 (Jin, Chen et al. 1998).[14][15] 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.[8][9] The alternative splice forms of ssTnT play a role in skeletal muscle adaptation in physiologic and pathological conditions.[16] Alternative splicing at alternative acceptor sites of intron 5 generates a single amino acid difference in the N-terminal region of ssTnT,[15] 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.[17][18] Truncation of the ssTnT polypeptide chain by the E180X mutation deletes the tropomyosin-binding site 2[19] 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.[20]
Tnnt1 gene targeted mouse studies reproduced the myopathic phenotypes of ANM.[21][22] 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 (Wei, Lu et al. 2014).[21]
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.[23] A Dutch patient with compound heterozygous TNNT1 gene mutations that cause exon 8 and exon 14 deletions also presents nemaline myopathy phenotypes.[24] 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.[25]
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.[19][26] The L203 truncation deletes the binding sites for TnI and TnC but preserves both tropomyosin-binding sites 1 and 2.[19] It remains to be investigated 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,[16] suggesting that structural modification of TnT in the myofilaments may contribute to adaptation to abnormalities in neuronal activation.
^"Human PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
^"Mouse PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
^Samson F, de Jong PJ, Trask BJ, Koza-Taylor P, Speer MC, Potter T, Roses AD, Gilbert JR (Aug 1992). "Assignment of the human slow skeletal troponin T gene to 19q13.4 using somatic cell hybrids and fluorescence in situ hybridization analysis". Genomics. 13 (4): 1374–5. doi:10.1016/0888-7543(92)90077-6. PMID1505979.
^Huang QQ, Jin JP (Dec 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". Journal of Molecular Evolution. 49 (6): 780–8. Bibcode:1999JMolE..49..780H. doi:10.1007/pl00006600. PMID10594179. S2CID9839814.
^Marra JD, Engelstad KE, Ankala A, Tanji K, Dastgir J, De Vivo DC, Coffee B, Chiriboga CA (May 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): 767–72. doi:10.1002/mus.24528. PMID25430424. S2CID29391961.
^Abdulhaq UN, Daana M, Dor T, Fellig Y, Eylon S, Schuelke M, Shaag A, Elpeleg O, Edvardson S (2015). "Nemaline body myopathy caused by a novel mutation in Troponin T1 (TNNT1)". Muscle Nerve. 53 (4): 564–9. doi:10.1002/mus.24885. PMID26296490. S2CID41713544.
^Amarasinghe C, Jin JP (Jun 2015). "N-Terminal Hypervariable Region of Muscle Type Isoforms of Troponin T Differentially Modulates the Affinity of Tropomyosin-Binding Site 1". Biochemistry. 54 (24): 3822–30. doi:10.1021/acs.biochem.5b00348. PMID26024675.
