A G86R mutation in the calcium-sensor protein GCAP1 alters regulation of retinal guanylyl cyclase and causes dominant cone-rod degeneration

• Published in: The Journal of biological chemistry Vol. 294; no. 10; pp. 3476 - 3488
• Main Authors: Peshenko, Igor V., Cideciyan, Artur V., Sumaroka, Alexander, Olshevskaya, Elena V., Scholten, Alexander, Abbas, Seher, Koch, Karl-Wilhelm, Jacobson, Samuel G., Dizhoor, Alexander M.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 08-03-2019; American Society for Biochemistry and Molecular Biology
• Subjects: Calcium - metabolism; calcium sensor; calcium-binding protein; Cell Death - genetics; Cone-Rod Dystrophies - enzymology; Cone-Rod Dystrophies - genetics; Cone-Rod Dystrophies - metabolism; Cone-Rod Dystrophies - pathology; cyclic GMP (cGMP); eye; guanylate cyclase (guanylyl cyclase); Guanylate Cyclase - metabolism; Guanylate Cyclase-Activating Proteins - chemistry; Guanylate Cyclase-Activating Proteins - genetics; Guanylate Cyclase-Activating Proteins - metabolism; GUCA1A; Humans; Models, Molecular; Mutation; photoreceptor; phototransduction; Protein Conformation, alpha-Helical; RetGC; Retina - enzymology; Retina - pathology; Retinal Cone Photoreceptor Cells - pathology; retinal degeneration; Retinal Rod Photoreceptor Cells - pathology; retinopathy; Signal Transduction; eye; retinal degeneration; calcium-binding protein; cyclic GMP (cGMP); calcium sensor; retinopathy; photoreceptor; guanylate cyclase (guanylyl cyclase); GUCA1A; phototransduction; RetGC
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.RA118.006180

The guanylyl cyclase-activating protein, GCAP1, activates photoreceptor membrane guanylyl cyclase (RetGC) in the light, when free Ca2+ concentrations decline, and decelerates the cyclase in the dark, when Ca2+ concentrations rise. Here, we report a novel mutation, G86R, in the GCAP1 (GUCA1A) gene in a family with a dominant retinopathy. The G86R substitution in a “hinge” region connecting EF-hand domains 2 and 3 in GCAP1 strongly interfered with its Ca2+-dependent activator-to-inhibitor conformational transition. The G86R-GCAP1 variant activated RetGC at low Ca2+ concentrations with higher affinity than did the WT GCAP1, but failed to decelerate the cyclase at the Ca2+ concentrations characteristic of dark-adapted photoreceptors. Ca2+-dependent increase in Trp94 fluorescence, indicative of the GCAP1 transition to its RetGC inhibiting state, was suppressed and shifted to a higher Ca2+ range. Conformational changes in G86R GCAP1 detectable by isothermal titration calorimetry (ITC) also became less sensitive to Ca2+, and the dose dependence of the G86R GCAP1–RetGC1 complex inhibition by retinal degeneration 3 (RD3) protein was shifted toward higher than normal concentrations. Our results indicate that the flexibility of the hinge region between EF-hands 2 and 3 is required for placing GCAP1-regulated Ca2+ sensitivity of the cyclase within the physiological range of intracellular Ca2+ at the expense of reducing GCAP1 affinity for the target enzyme. The disease-linked mutation of the hinge Gly86, leading to abnormally high affinity for the target enzyme and reduced Ca2+ sensitivity of GCAP1, is predicted to abnormally elevate cGMP production and Ca2+ influx in photoreceptors in the dark.