Proteostatic significance of helix O alanine residues in CLC channels

The voltage-gated chloride channels ClC-1 and ClC-2 are homodimeric structures essential for maintaining muscle excitability and tissue fluid homeostasis, respectively. Mutations in these channels disrupt protein homeostasis (proteostasis), leading to hereditary disorders such as myotonia congenita and leukodystrophy. Specifically, the substitution of highly conserved alanine residues within the transmembrane helix O, exemplified by ClC-1 (A531V) and ClC-2 (A500V), results in severe proteostatic defects characterized by reduced protein stability and impaired surface trafficking. However, the precise role of helix O in these pathological processes remains poorly understood. In this study, we investigated these conserved residues using biochemical and functional approaches. Our findings demonstrate that even subtle structural alterations at these critical sites significantly interfere with channel stability and membrane expression. This study highlights the critical contribution of helix O to proper CLC channel folding and endoplasmic reticulum (ER) quality control, providing deeper insights into the molecular mechanisms of CLC-related channelopathies.