Cadherin superfamily in intercellular junctions of podocytes in the kidney

在用简并引物RT-PCR寻找大鼠肾小球中表达的钙粘蛋白相关分子的过程中，我们分离到了编码大鼠人类FAT同源物的cDNA。FAT是钙粘蛋白超家族的一个新成员，类似于果蝇的抑癌脂肪。它是一个大的跨膜蛋白，有34个串联类钙粘素重复序列，5个类EGF重复序列，以及一个层粘连蛋白A-G结构域，表明它在细胞间黏附和细胞增殖中具有潜在的作用。为了定位脂肪并推测其在肾脏中的作用，我们用核糖核酸酶保护分析(RPA)、原位杂交(ISH)和免疫染色等方法检测了脂肪在生理和病理条件下在新生大鼠和成年大鼠肾脏中的表达。推导的氨基酸序列在大鼠和人类脂肪中有85%以上的同源性，特别是在C端的同源性为100%。制备了抗脂肪C末端的亲和纯化抗体。RPA在正常大鼠肾小球的RNA中显示明显的信号。免疫组织化学显示足细胞有脂肪表达。脂肪免疫染色结果与5.1.6的结果一致，后者是针对裂隙隔的特异性抗体。在PAN肾病中，RPA、ISH和免疫组织化学染色显示肾小球脂肪表达明显增加。在新生大鼠肾脏，从S形体到成熟肾小球的整个发育过程中，足细胞内均可观察到脂肪的存在，提示脂肪是足细胞细胞间连接的基本成分之一。

During a search for cadherin-related molecules expressed in rat glomeruli by PT-PCR with degenerate primers, we have isolated cDNA encoding a rat homologue of human FAT.FAT, a new member of the cadherin superfamily, resembles the Drosophila tumor suppressor fat. It is a large transmembrane protein with 34 tandem cadherin-like repeats, five EGF-like repeats, and a laminin A-G domain, indicating its potential role in cell-cell adhesion and cell proliferation. To localize FAT and to speculate its role in the kidney, we examined its expression in the rat neonatal and adult kidney in the physiological and pathological conditons by using ribonuclease protection assay (RPA), in situ hybridization(ISH) and immunostaining.Comparison of the deduced amino acid sequences showed more than 85% identical residues in rat and human FAT, especially, 100% identical in C-terminus of FAT of the both species. Affinity-purified antibody against FAT C-terminus was prepared. RPA revealed significant signals in RNA from glomeruli of the normal rat kidney. By ISH, FAT expression was observed in podocytes. Immunostaining for FAT was coincident with that of 5.1.6, which is antibody specific for slit diaphragm. In PAN nephrosis, conspicuous increase in FAT expression in glomeruli was shown by RPA, ISH, and immunostaining. In the neonatal kidney, FAT was observed in podocytes throughout the development from S-shaped body to mature glomeruli.The above findings suggest that FAT is one of basic constitutents of intercellular junctions of podocytes.