Molday L L, Hicks D, Sauer C G, Weber B H, Molday R S
Department of Biochemistry and Molecular Biology, University of British Columbia, 2146 Health Sciences Mall, Vancouver, British Columbia V6T 1Z3, Canada.
Invest Ophthalmol Vis Sci. 2001 Mar;42(3):816-25.
To examine the biochemical properties, cell expression, and localization of RS1, the product of the gene responsible for X-linked juvenile retinoschisis.
Rs1h mRNA expression was measured from the eyes of wild-type and rd/rd mice by Northern blot analysis and reverse transcription-polymerase chain reaction (RT-PCR). Specific antibodies raised against the N terminus of RS1 were used as probes to examine the properties and distribution of RS1 in retina, retinal cell cultures, and transfected COS-1 cells by Western blot analysis and immunofluorescence microscopy.
Rs1h mRNA expression was detected in the retina of postnatal day (P)11 and adult CD1 mice, but not homozygous rd/rd mice by Northern blot analysis. However, Rs1h expression was detected in rd/rd mice by RT-PCR. RS1 migrated as a single 24-kDa polypeptide under disulfide-reducing conditions and a larger complex (>95 kDa) under nonreducing conditions in the membrane fraction of retinal tissue homogenates and transfected COS-1 cells. RS1 antibodies specifically stained rod and cone photoreceptors and most bipolar cells, but not Müller cells, ganglion cells, or the inner limiting membrane of adult and developing retina as revealed in double-labeling studies. RS1 antibodies also labeled retinal bipolar cells of photoreceptorless mice and retinal bipolar cells grown in cell culture.
RS1 is expressed and assembled in photoreceptors of the outer retina and bipolar cells of the inner retina as a disulfide-linked oligomeric protein complex. The secreted complex associates with the surface of these cells, where it may function as a cell adhesion protein to maintain the integrity of the central and peripheral retina.
研究与X连锁青少年视网膜劈裂症相关基因的产物RS1的生化特性、细胞表达及定位。
通过Northern印迹分析和逆转录-聚合酶链反应（RT-PCR）检测野生型和rd/rd小鼠眼中Rs1h mRNA的表达。用针对RS1 N端产生的特异性抗体作为探针，通过蛋白质免疫印迹分析和免疫荧光显微镜检查RS1在视网膜、视网膜细胞培养物和转染的COS-1细胞中的特性和分布。
通过Northern印迹分析，在出生后第11天（P11）的视网膜和成年CD1小鼠的视网膜中检测到Rs1h mRNA表达，但在纯合rd/rd小鼠中未检测到。然而，通过RT-PCR在rd/rd小鼠中检测到Rs1h表达。在视网膜组织匀浆和转染的COS-1细胞的膜部分中，RS1在二硫键还原条件下迁移为单一的24 kDa多肽，在非还原条件下迁移为更大的复合物（>95 kDa）。双重标记研究显示，RS1抗体特异性地标记视杆和视锥光感受器以及大多数双极细胞，但不标记Müller细胞、神经节细胞或成年和发育中视网膜的内界膜。RS1抗体也标记无感光细胞小鼠的视网膜双极细胞和细胞培养中生长的视网膜双极细胞。
RS1在外视网膜的光感受器和内视网膜的双极细胞中表达并组装为二硫键连接的寡聚蛋白复合物。分泌的复合物与这些细胞的表面结合，在那里它可能作为细胞粘附蛋白发挥作用，以维持中央和周边视网膜的完整性。
