Feng Han-Zhong, Strauss Kevin A, Jin Jian-Ping
Department of Physiology and Biophysics, University of Illinois at Chicago, IL, USA.
Clinic for Special Children, Gordonville, PA, USA.
FEBS J. 2025 Jun 26. doi: 10.1111/febs.70165.
A nonsense mutation in codon Glu of the TNNT1 gene, which encodes the slow skeletal muscle isoform of troponin T (ssTnT), causes a recessively inherited myopathy (the Amish Nemaline Myopathy, ANM). A ssTnT knockout (ssTnT-KO) mouse model produced the loss of ssTnT function phenotypes of ANM with slow fiber atrophy and decreased fatigue resistance of soleus muscle. We further developed a Tnnt1 p.Glu180* knock-in (ANM-KI) mouse model to precisely mimic the human mutation. In addition to reproducing the loss of function phenotypes, ANM-KI mice exhibit more severe myopathy than that of ssTnT-KO mice. Compared with wild-type controls, ANM-KI and ssTnT-KO soleus muscles show different changes in gene expression profiles, of which gene ontology analysis indicated inflammatory activation in ANM-KI soleus muscle. The mutant Tnnt1 mRNA was readily detectable in ANM-KI soleus muscle. However, the truncated ssTnT fragment cannot be detected in western blot, indicating its very low level due to the active proteolytic clearance of non-myofilament-incorporated TnT in muscle cells. Nonetheless, the more severe myopathic impacts of the ANM-KI allele with more fiber number loss and muscle activity/injury-caused hypertrophy support a potent cytotoxicity of the ssTnT fragment, as shown in previous cell culture studies, which is further supported by activity-dependent and age-progressing myopathy with more active regeneration. The notion that non-myofilament-incorporated ssTnT fragments may potentially contribute to the pathogenesis and progression of myopathy merits further investigation.
TNNT1基因密码子Glu发生无义突变，该基因编码肌钙蛋白T的慢肌亚型（ssTnT），会导致一种隐性遗传的肌病（阿米什人杆状体肌病，ANM）。一种ssTnT基因敲除（ssTnT-KO）小鼠模型产生了ANM的ssTnT功能缺失表型，伴有慢肌纤维萎缩和比目鱼肌疲劳抗性降低。我们进一步构建了一个Tnnt1 p.Glu180*基因敲入（ANM-KI）小鼠模型，以精确模拟人类突变。除了重现功能缺失表型外，ANM-KI小鼠表现出比ssTnT-KO小鼠更严重的肌病。与野生型对照相比，ANM-KI和ssTnT-KO比目鱼肌在基因表达谱上有不同变化，基因本体分析表明ANM-KI比目鱼肌存在炎症激活。在ANM-KI比目鱼肌中很容易检测到突变的Tnnt1 mRNA。然而，在蛋白质印迹中无法检测到截短的ssTnT片段，这表明由于肌肉细胞中非肌丝结合的TnT被活跃的蛋白水解清除，其水平非常低。尽管如此，ANM-KI等位基因更严重的肌病影响，伴随着更多肌纤维数量减少以及肌肉活动/损伤引起的肥大，支持了ssTnT片段具有强大的细胞毒性，正如先前细胞培养研究所显示的那样，这进一步得到了活动依赖性和年龄进展性肌病以及更活跃再生的支持。非肌丝结合的ssTnT片段可能潜在地导致肌病发病机制和进展这一观点值得进一步研究。
