Interphotoreceptor retinoid-binding protein (IRBP): Expression in the adult and developingXenopus retina

Abstract

Apposition of the neural retina and pigment epithelium is critical to photoreceptor development and function. Interphotoreceptor retinoid-binding protein (IRBP) is a major component of the extracellular matrix separating these epithelia in the African clawed frog Xenopus laevis (Gonzalez-Fernandez et al., [1993], J. Cell Sci. 105:7-21). In the adult retina, IRBP appears to mediate the transport of hydrophobic molecules, particularly retinoids and fatty acids, within the hydrophilic extracellular domain. In this paper, we compare the distribution of IRBP and its mRNA in adult and embryonic Xenopus retina. Xenopus IRBP antisense RNA, labeled with tritium or digoxigenin, was used for in situ hybridizaton studies. For immunohistochemistry, we used an antiserum against Xenopus IRBP expressed in Escherichia coli. In the adult, we found that IRBP is synthesized at similar levels by both rods and cones. The protein is restricted to the interphotoreceptor matrix, with lesser amounts in the pigment epithelial cytoplasm. In the embryo, expression of the mRNA for IRBP is restricted to the central retina, where photoreceptor differentiation has taken place. By contrast, the protein is distributed throughout the embryonic subretinal space. Therefore, the presence of IRBP precedes photoreceptor differentiation. In summary, IRBP is synthesized by both rods and cones and may be internalized by the pigment epithelium. In the embryo, IRBP is synthesized by the central retina and diffuses through the matrix, reaching the undifferentiated peripheral retina. In view of its ligand-binding properties, diffusion of IRBP may provide the peripheral neural retina with a vehicle to transport retinoids and docosahexaenoic acid (molecules critical to normal retinal development) from the pigment epithelium.

Fourth module of Xenopus interphotoreceptor retinoid-binding protein: activity in retinoid transfer between the retinal pigment epithelium and rod photoreceptors

“…The posterior hemisphere was embedded in Lowicryl resin (K4M, Polysciences) at 4 Њ C. Seventy-nm sections were incubated in a 1:200 dilution of anti-X4IRBP serum or preimmune control serum. 35 Sections were treated with protein A (Sigma) labeled with 10 nm colloidal gold. The grids were then fixed in 2% glutaraldehyde and 2% OsO 4 , counterstained with 5% uranyl acetate in 50% acetone and Reynold's lead citrate, and examined in a JEOL 100CX transmission electron microscope.…”

Goldfish cones secrete a two-repeat interphotoreceptor retinoid-binding protein

“…Although rods make up the majority of photoreceptors, a clear association of grains with their inner segments was never convincing. In contrast, the inner segments of both Xenopus rods and cones hybridize with antisense IRBP RNA (Hessler et al , 1995.…”

“…The blot was blocked for 2 h in rinsing solution containing 3% bovine serum albumin (BSA, fraction V, Sigma) and 3% powdered milk. This was foilowed by incubating 4 h in rabbit antiXenopus IRBP serum (Hessler et al 1995) at a 1:1,500 dilution in TBS containing 3% BSA. The blot was washed in rinsing solution and incubated with a horseradish-peroxidaseconjugated donkey antirabbit IgG at a 1:2,000 dilution in blocking solution and detected by chemiluminescence (Amersham, Arlington Heights,IL).…”

“…This is accomplished by interphotoreceptor retinoid-binding protein (IRBP), a large (145 kDa in bovine) glycolipoprotein, the major protein component of the matrix. IRBP is secreted into the matrix by rods and cones (GonzalezFernandez et al 1984(GonzalezFernandez et al , 1992Hollyfield et al 1985a;Van Veen et al 1986;Porrello et al 1991;Carter-Dawson and Burroughs 1992;Yokoyama et al 1992;Hessler et al 1995) and removed from the matrix by phagocytic and nonphagocytic endocytic pathways (Hollyfield et al 1985a;Payman et al 1994). IRBP facilitates the diffusion of retinoids, promotes the release of 11-cis retinal from the pigmented epithelium and protects the isomeric and oxidation state of retinol (reviewed by Carlson and Bok 1992;Crouch 1992;Pepperberg et al 1993).…”

Enzymatic heterogeneity of bovine retinal pigment epithelial cells in vivo and in vitro

“…The membrane-associated proteases aminopeptidases A and M and dipeptidylpeptidase IV were localized because they may be involved in the turnover of proteins in the subretinal space and the interphotoreceptor matrix. These proteins (e.g., interphotoreceptor retinoid-binding protein) are metabolized predominantly by the RPE cells [20]. Alkaline phosphatase in the RPE appears to be necessary for the transport of solutes between choroid capillaries and subretinal space [11].…”