Proteoglycans in the mouse interphotoreceptor matrix. VI. Evidence for photoreceptor synthesis of chondroitin sulfate proteoglycan using genetically fractionated retinas.
To determine the role of photoreceptors in the synthesis of chondroitin sulfate proteoglycan (CS-PG) present in the interphotoreceptor matrix (IPM), 35SO4(2-) was used as a tracer for comparison of proteoglycans synthesized in vitro in the absence of the pigment epithelium by normal retinas and retinas from retinal degeneration (rd) mice at stages before and after photoreceptor degeneration. Isolated retinas from 10 day post-partum (P-10) pups, adult normal mice (C57BL/6J ++/++) and retinal degeneration mice (C57BL/6J rdle/rdle) were incubated for 7 hr with 35SO4(2-) to label newly synthesized sulfated proteoglycans. At P-10, rd retinas have not undergone extensive photoreceptor degeneration, whereas in the adult retinas from this strain, only a few cone photoreceptors remain. At the termination of the labeling period, proteoglycans in the incubation medium and those remaining in guanidine hydrochloride (GuHCl) extracts of the retina were analysed separately and identified by their susceptibility to enzymatic or nitrous acid depolymerization. At P-10, no significant differences were observed in the types or sizes of newly synthesized proteoglycan in normal and rd retinas. Medium samples from P-10 retinas contained near equal amounts of 35S-labeled CS-PG and heparan sulfate proteoglycan (HS-PG), while in GuHCl extracts, approximately 90% of the 35SO4(2-) was incorporated into HS-PG, with the remainder found in CS-PG. Comparisons of adult tissue revealed a divergence of proteoglycan synthesis profiles. Retinas from normal adults label predominantly CS-PG. [35S]proteoglycan from normal retina incubation medium was approximately 96% CS-PG, and GuHCl extracts were about 73% CS-PG. From adult rd retinas these values were 18 and 10%, respectively. Per retina, this shows the rd retinas labeling less than 4% of the medium CS-PG, and about 50% of the GuHCl extractable CS-PG compared to normal retinas. Labeled HS-PG comprised about 28% of the normal retina GuHCl extracts, but was not detected in the incubation medium. In contrast, HS-PG synthesis accounted for about 76% of the medium proteoglycan label, and about 85% of the extracted proteoglycan in the adult rd retina. In fact, 35SO4(2-) labeling of HS-PG in the rd retina GuHCl extracts exceeded by 1000% the level observed in normal retina extracts on a per retina basis. Retinas from both strains incorporate significant amounts of 35SO4(2-) into proteins with rd achieving higher specific activity. IRBP was identified as a 35SO4(2-) labeled protein by immunoadsorption from aliquots of the incubation medium.(ABSTRACT TRUNCATED AT 400 WORDS)
Landers
R A
RA
Cullen Eye Institute, Baylor College of Medicine, Houston, TX 77030.
Hollyfield
J G
JG
D000818:Animals; D011508:Chondroitin Sulfate Proteoglycans / Q000096:biosynthesis*; D002850:Chromatography, Gel; D005136:Eye Proteins / Q000096:biosynthesis; D005260:Female; D006497:Heparan Sulfate / Q000096:biosynthesis; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D010786:Photoreceptor Cells / Q000378:metabolism*; D012160:Retina / Q000473:pathology; D012162:Retinal Degeneration / Q000378:metabolism* / Q000473:pathology; D012177:Retinol-Binding Proteins / Q000096:biosynthesis; D013462:Sulfur Radioisotopes; D000099214:Retinol-Binding Proteins, Interstitial
D011508:Chondroitin Sulfate Proteoglycans; D005136:Eye Proteins; D006497:Heparan Sulfate; D012177:Retinol-Binding Proteins; D013462:Sulfur Radioisotopes; D000099214:Retinol-Binding Proteins, Interstitial