Proteomic Identification of Unique Photoreceptor Disc Components Reveals the Presence of PRCD, a Protein Linked to Retinal Degeneration

Abstract

Visual signal transduction takes place on the surface of flat membrane vesicles called photoreceptor discs, which reside inside the light-sensitive outer segment organelle of vertebrate photoreceptor cells. While biochemical studies have indicated that discs are built with a handful of highly specialized proteins, proteomic studies have yielded databases consisting of hundreds of entries. We addressed this controversy by employing protein correlation profiling which allows identification of unique components of organelles that can be fractionated but not purified to absolute homogeneity. We subjected discs to sequential steps of fractionation and identified the relative amounts of proteins in each fraction by label-free quantitative mass spectrometry. This analysis demonstrated that the photoreceptor disc proteome contains only eleven components, which satisfy the hallmark criterion for being unique disc-resident components: the retention of a constant molar ratio among themselves across fractionation steps. Remarkably, one of them is PRCD, a protein whose mutations have been shown to cause blindness, yet cellular localization remained completely unknown. Identification of PRCD as a novel disc-specific protein facilitates understanding its functional role and the pathobiological significance of its mutations. Our study provides a striking example how protein correlation profiling allows a distinction between constitutive components of cellular organelles and their inevitable contaminants.

Palmitoylation of Progressive Rod-Cone Degeneration (PRCD) Regulates Protein Stability and Localization

“…Association of PRCD with Membranes Is Not Dependent on Palmitoylation-A previous study revealed that PRCD is one of 11 proteins exclusively present in the photoreceptor disc membrane (16). We wanted to investigate the contribution of protein palmitoylation in the membrane association of PRCD.…”

“…A proteomic study demonstrated that PRCD is one of 11 proteins exclusively present in the photoreceptor outer segment (OS) disc membrane (16). The presence of PRCD within the OS disc membranes suggests that it plays an important role in photoreceptor function and/or maintenance.…”

Interaction of 4.1G and cGMP-gated channels in rod photoreceptor outer segments

“…The mass spectrometry analysis performed on samples immunoprecipitated with the CNGA1 and CNGB1 antibodies failed to detect ankyrin G peptides. In fact, ankyrin G has not been found in massspectrometric-based proteomic studies of photoreceptor outer segments (Kiel et al, 2011;Kwok et al, 2008;Reidel et al, 2011;Skiba et al, 2013). Considering the size of this protein it is surprising that these proteomics studies have failed to detect peptides from ankyrin G in outer segment preparations.…”

In vivo optophysiology reveals that G-protein activation triggers osmotic swelling and increased light scattering of rod photoreceptors

“…Mouse rod outer segments (OS) are in effect genetically manipulable, femtoliter test tubes whose protein constituents (15,16), as well as their biochemical reactions (17)(18)(19) and light-stimulated translocations (20)(21)(22), are well known and have been measured primarily with ex vivo methods. Advances in imaging technology have now made it possible to achieve subcellular-resolution imaging of the mouse retina in vivo (23)(24)(25)(26).…”