Signal Transduction
Two clusters of surface-exposed amino acid residues enable high-affinity binding of retinal degeneration-3 (RD3) protein to retinal guanylyl cyclase
Retinal degeneration-3 (RD3) protein protects photoreceptors from degeneration by preventing retinal guanylyl cyclase (RetGC) activation via calcium-sensing guanylyl cyclase–activating proteins (GCAP), and RD3 truncation causes severe congenital blindness in humans and other animals. The three-dimensional structure of RD3 has recently been established, but the molecular mechanisms of its inhibitory binding to RetGC remain unclear. Here, we report the results of probing 133 surface-exposed residues in RD3 by single substitutions and deletions to identify side chains that are critical for the inhibitory binding of RD3 to RetGC.Retinal guanylyl cyclase activation by calcium sensor proteins mediates photoreceptor degeneration in an rd3 mouse model of congenital human blindness
Deficiency of RD3 (retinal degeneration 3) protein causes recessive blindness and photoreceptor degeneration in humans and in the rd3 mouse strain, but the disease mechanism is unclear. Here, we present evidence that RD3 protects photoreceptors from degeneration by competing with guanylyl cyclase-activating proteins (GCAPs), which are calcium sensor proteins for retinal membrane guanylyl cyclase (RetGC). RetGC activity in rd3/rd3 retinas was drastically reduced but stimulated by the endogenous GCAPs at low Ca2+ concentrations.A G86R mutation in the calcium-sensor protein GCAP1 alters regulation of retinal guanylyl cyclase and causes dominant cone-rod degeneration
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.Structure of Guanylyl Cyclase Activator Protein 1 (GCAP1) Mutant V77E in a Ca2+-free/Mg2+-bound Activator State
GCAP1, a member of the neuronal calcium sensor subclass of the calmodulin superfamily, confers Ca2+-sensitive activation of retinal guanylyl cyclase 1 (RetGC1). We present NMR resonance assignments, residual dipolar coupling data, functional analysis, and a structural model of GCAP1 mutant (GCAP1V77E) in the Ca2+-free/Mg2+-bound state. NMR chemical shifts and residual dipolar coupling data reveal Ca2+-dependent differences for residues 170–174. An NMR-derived model of GCAP1V77E contains Mg2+ bound at EF2 and looks similar to Ca2+ saturated GCAP1 (root mean square deviations = 2.0 Å).Dimerization Domain of Retinal Membrane Guanylyl Cyclase 1 (RetGC1) Is an Essential Part of Guanylyl Cyclase-activating Protein (GCAP) Binding Interface
Background: GCAPs regulate photoreceptor guanylyl cyclase RetGC1 but not hormone receptor guanylyl cyclase NPRA.Results: Mutations in RetGC1 dimerization domain disrupt GCAP1 and GCAP2 binding.Conclusion: Met823 in dimerization domain strongly contributes to its specificity in forming GCAP binding interface.Significance: Congenital blindness-causing mutation in the neighboring residue prohibits GCAP binding.
