Molecular pathogenicity of disease-associated mutations in cone CNG channel subunits

Chunming Liu

doi:10.7273/000005863

Located at the outer segment membrane of photoreceptor cells, cyclic nucleotide-gated CNG channels are a critical step in the phototransduction cascade by which light stimulation is converted into electrical responses that can be processed by the central nervous system. The abundance and proper functioning of CNG channels are critical for photoreceptor survival. Multiple mutations in genes encoding CNG channel subunits have been identified in patients with inherited retinal degenerative diseases, a genetically heterogeneous group of disorders characterized by loss of rod, cone, or both photoreceptor cells. The resulting blindness is one of the major causes of disability. However, the pathogenic mechanisms arising from these mutations have remained largely undetermined. To address this question, we have focused on cone specific CNG channels and functionally characterized three mutations associated with severe progressive cone dystrophy. Complex effects, including altered channel-gating properties, impaired channel subunit processing and maturation, and disrupted plasma membrane localization, were observed with these mutations. These results provide evidence for the pathogenecity of the mutations in humans, and suggest that changes in channel activity and/or localization may ultimately lead to the reported clinical features. To further identify the possible cellular and molecular pathways by which CNG channel mutations initiate photoreceptor degeneration, two achromatopsia-associated, gain-of-function mutations in CNGB3 were studied using heterologous expression in photoreceptor derived 661W cell. Mutant channels exhibited increased apparent affinity for CPT-cGMP compared to wild-type channels. Increased cytotoxicity induced by a low concentration of CPT-cGMP was observed in mutant channel-expressing 661W cells. The elevation in cytotoxicity was found to be dependent on the presence of extracellular calcium. The increased susceptibility to cell death could be rescued by applying CNG channel blockers L-cis-diltiazem or tetracaine. These results suggest that cytotoxicity in this model depends on calcium entry through hyperactive CNG channels, supporting a Ca2+-overload hypothesis. Our results are the first to show a direct connection between disease-associated mutations in cone CNG channel subunits, altered CNG channel-activation properties, and photoreceptor cytotoxicity.

Molecular pathogenicity of disease-associated mutations in cone CNG channel subunits

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