Abstract
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. RetGC activity completely failed to accelerate in rd3/rd3GCAPs-/- hybrid photoreceptors, whose photoresponses remained drastically suppressed compared with the WT. However, ∼70% of the hybrid rd3/rd3GCAPs-/- photoreceptors survived past 6 months, in stark contrast to <5% in the nonhybrid rd3/rd3 retinas. GFP-tagged human RD3 inhibited GCAP-dependent activation of RetGC in vitro similarly to the untagged RD3. When transgenically expressed in rd3/rd3 mouse retinas under control of the rhodopsin promoter, the RD3GFP construct increased RetGC levels to near normal levels, restored dark-adapted photoresponses, and rescued rods from degeneration. The fluorescence of RD3GFP in rd3/rd3RD3GFP+ retinas was mostly restricted to the rod photoreceptor inner segments, whereas GCAP1 immunofluorescence was concentrated predominantly in the outer segment. However, RD3GFP became distributed to the outer segments when bred into a GCAPs-/- genetic background. These results support the hypothesis that an essential biological function of RD3 is competition with GCAPs that inhibits premature cyclase activation in the inner segment. Our findings also indicate that the fast rate of degeneration in RD3-deficient photoreceptors results from the lack of this inhibition.
Highlights
Deficiency of RD3 protein causes recessive blindness and photoreceptor degeneration in humans and in the rd3 mouse strain, but the disease mechanism is unclear
Our results indicate that prevention of retinal membrane guanylyl cyclase (RetGC) activation by guanylyl cyclaseactivating proteins (GCAPs) in the inner segment is a critical biological function of RD3 in photoreceptor physiology
The role of RD3 in maintaining the RetGC levels is most critical for the function of photoreceptors—the phototransduction becomes inefficient when the cGMP synthesis is impaired, forcing most of the cGMP gated channels that would be normally open in the dark to become closed [17, 22, 26] (Fig. 1B)
Summary
Production of cGMP in photoreceptors by retinal membrane RetGC2 (isozymes RetGC1, GUCY2D) and RetGC2 (GUCY2F) [1,2,3] imparts light sensitivity to vertebrate photoreceptors by. The authors declare that they have no conflicts of interest with the contents of this article.
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