Abstract：
Autophagy phenomenon in the cell maintains proteostasis balance by eliminating damaged organelles and protein aggregates. Imbalance in autophagic flux may cause accumulation of protein aggregates in various neurodegenerative disorders. Regulation of autophagy by either calcium or chaperone play a key role in the removal of protein aggregates from the cell. The neuromuscular rare genetic disorder, GNE Myopathy, is characterized by accumulation of rimmed vacuoles having protein aggregates of β-amyloid and tau that may result from altered autophagic flux. In the present study, the autophagic flux was deciphered in HEK cell-based model for GNE Myopathy harbouring GNE mutations of Indian origin. The refolding activity of HSP70 chaperone was found to be reduced in GNE mutant cells compared to wild type controls. The autophagic markers LC3II/I ratio was altered with increased number of autophagosome formation in GNE mutant cells compared to wild type cells. The cytosolic calcium levels were also increased in GNE mutant cells of Indian origin. Interestingly, treatment of GNE mutant cells with HSP70 activator, BGP-15, restored the expression and refolding activity of HSP70 along with autophagosome formation. Treatment with calcium chelator, BAPTA-AM restored the cytoplasmic calcium levels and autophagosome formation but not LC3II/I ratio significantly. Our study provides insights towards GNE mutation specific response for autophagy regulation and opens up a therapeutic advancement area in calcium signalling and HSP70 function for GNE related Myopathy.
摘要：
细胞中的自噬现象通过消除受损的细胞器和蛋白质聚集体来维持蛋白质平衡。自噬通量的失衡可能导致各种神经退行性疾病中蛋白质聚集体的积累。钙或伴侣对自噬的调节在从细胞中去除蛋白质聚集体中起关键作用。神经肌肉罕见的遗传病，GNE肌病,其特征在于具有β-淀粉样蛋白和tau蛋白聚集体的有边液泡的积累，这可能是由改变的自噬通量引起的。在本研究中,在基于HEK细胞的GNE肌病模型中破译了自噬通量，该模型具有印度裔GNE突变。发现与野生型对照相比,在GNE突变细胞中HSP70伴侣的重折叠活性降低。与野生型细胞相比,自噬标记物LC3II/I比率随着GNE突变细胞中自噬体形成数量的增加而改变。印度裔GNE突变细胞的胞浆钙水平也增加。有趣的是,用HSP70激活剂处理GNE突变细胞，BGP-15恢复了HSP70的表达和重折叠活性，并形成自噬体。用钙螯合剂治疗，BAPTA-AM恢复了细胞质钙水平和自噬小体形成，但没有显着恢复LC3II/I比率。我们的研究为自噬调节的GNE突变特异性反应提供了见解，并为GNE相关肌病的钙信号和HSP70功能开辟了治疗进展领域。
