Ochrietor Judith D, Moroz Tatiana P, van Ekeris Leslie, Clamp Michael F, Jefferson Stephanie C, deCarvalho Ana C, Fadool James M, Wistow Graeme, Muramatsu Takashi, Linser Paul J
Whitney Laboratory of the University of Florida, St. Augustine, Florida 32080, USA.
Invest Ophthalmol Vis Sci. 2003 Sep;44(9):4086-96. doi: 10.1167/iovs.02-0995.
5A11/Basigin has recently been identified as a critical glycoprotein for full maturity and function of the mouse retina. However, the biological function of 5A11/Basigin has yet to be determined. Previous reports indicate the presence of multiple 5A11/Basigin polypeptides within the retina. Therefore, in an effort to determine the function of 5A11/Basigin, the molecular diversity of its expression was evaluated.
Northern blot and immunoblot techniques were used to evaluate the number of forms of 5A11/Basigin in the mouse retina. cDNA cloning, using a mouse retina library or RT-PCR from rat, chicken, zebrafish, and human retina, was performed to determine the sequence of 5A11/Basigin transcripts. A peptide was generated, based on the deduced amino acid sequence, for subsequent antibody production. Localization of 5A11/Basigin expression was evaluated by immunoblot, immunohistochemistry, and real-time RT-PCR.
Two 5A11/Basigin transcripts of approximately 1.5 kb and approximately 1.8 kb, which correspond to glycosylated proteins of approximately 45 and approximately 55 kDa, respectively, were identified in mouse retina. The shorter form was previously cloned. However, the longer form, a splice variant of mouse 5A11/Basigin, is a member of the immunoglobulin gene superfamily and has been named 5A11/Basigin-2. Homologous transcripts were also cloned from rat, chicken, zebrafish, and human retina. 5A11/Basigin-2 expression was limited to the retina, specifically to photoreceptor cells, where it appeared to be most concentrated in the inner segments.
The specific and limited expression of 5A11/Basigin-2 explicitly within photoreceptor cells implies that this glycoprotein plays a fundamental role within the retina. However, its role remains to be determined.
5A11/基底膜蛋白最近被确定为小鼠视网膜完全成熟和发挥功能所必需的一种关键糖蛋白。然而，5A11/基底膜蛋白的生物学功能尚未明确。此前有报道称视网膜内存在多种5A11/基底膜蛋白多肽。因此，为了确定5A11/基底膜蛋白的功能，对其表达的分子多样性进行了评估。
采用Northern印迹法和免疫印迹技术评估小鼠视网膜中5A11/基底膜蛋白的形式数量。利用小鼠视网膜文库或从大鼠、鸡、斑马鱼和人类视网膜进行RT-PCR进行cDNA克隆，以确定5A11/基底膜蛋白转录本的序列。根据推导的氨基酸序列生成一种肽，用于后续抗体的制备。通过免疫印迹、免疫组织化学和实时RT-PCR评估5A11/基底膜蛋白表达的定位。
在小鼠视网膜中鉴定出两种5A11/基底膜蛋白转录本，大小分别约为1.5 kb和1.8 kb，分别对应于约45 kDa和约55 kDa的糖基化蛋白。较短的形式先前已被克隆。然而，较长的形式是小鼠5A11/基底膜蛋白的剪接变体，属于免疫球蛋白基因超家族成员，已被命名为5A11/基底膜蛋白-2。也从大鼠、鸡、斑马鱼和人类视网膜中克隆出了同源转录本。5A11/基底膜蛋白-2的表达仅限于视网膜，特别是光感受器细胞，在那里它似乎在内段最为集中。
5A11/基底膜蛋白-2在光感受器细胞内特异性且有限的表达表明，这种糖蛋白在视网膜中发挥着重要作用。然而，其作用仍有待确定。
