shRNA knockdown ofguanylate cyclase 2eorcyclic nucleotide gated channel alpha 1increases photoreceptor survival in a cgmpphosphodiesterasemouse model of retinitis pigmentosa

In vertebrate rods, dark and light conditions produce changes in guanosine 3',5'-cyclic monophosphate (cGMP) and calcium (Ca ²⁺) levels, which are regulated by the opposing function of several proteins. During the recovery of a bright flash, guanylate cyclase (GUCY) helps raise cGMP to levels that open cGMP-gated calcium sodium channels (CNG) to increase Na ⁺ and Ca ²⁺ influx in the outer segment. In contrast, light activates cGMP phosphodiesterase 6 (PDE6) causing rapid hydrolysis of cGMP, CNG closure, and reduced Na ⁺ and Ca ²⁺ levels. InPde6bmouse models of retinitis pigmentosa (RP), photoreceptor death is preceded by abnormally high cGMP and Ca ²⁺ levels, likely because of continued synthesis of cGMP by guanylate cyclases and unregulated influx of Ca ²⁺ to toxic levels through CNG channels. To reverse the effects ofPde6bloss of function, we employed an shRNA knockdown approach to reduce the expression ofGucy2eorCnga1inPde6b ^H620Q photoreceptors prior to degeneration.Gucy2e- orCnga1-shRNA lentiviral-mediated knockdown GUCY2E and CNGA1 expression increase visual function and photoreceptor survival inPde6b ^H620Q mice. We demonstrated that effective knockdown ofGUCY2EandCNGA1expression to counteract loss of PDE6 function may develop into a valuable approach for treating some patients with RP.