PATHOGENESIS OF FAMILIAL JUVENILE NEPHRONOPHTHISIS

这项提案的长期目标是了解家族性青少年肾单位营养不良（NPH）-儿童肾衰竭的常见遗传原因-发病机制的细胞缺陷的性质。 虽然临床和组织学观察表明NPH是由于正常排泄小管功能丧失所致，但该病的发病机制仍不清楚。 最近，在大多数NPH病例中突变的人类基因被分离出来，并确定了其编码蛋白质（“肾囊素”）的序列。 肾囊蛋白的生化功能的线索是已知介导特异性蛋白质-蛋白质相互作用的“Src同源3”（SH 3）结构域的存在。 对肾囊素的进一步研究将为NPH的发病机制提供新的认识。 在初步研究中，编码小鼠肾囊蛋白的cDNA已被分离，并已在植入后小鼠胚胎发生过程中和包括肾脏在内的各种成人组织中检测到转录本。 通过鉴定酪氨酸激酶底物p130 Cas作为肾囊蛋白SH 3结构域结合的蛋白质，并通过显示肾囊蛋白定位于极化上皮细胞的侧膜，获得了关于肾囊蛋白的生化功能的新信息。 这些观察结果导致我们的一般假设，即肾囊蛋白在肾小管上皮的形态发生和/或维持中起作用，需要与pl 30 Cas和参与细胞粘附相互作用的其他蛋白质的特异性相互作用。 为了验证和扩展这一假设，提出了具体的目标：1）确定肾囊蛋白在发育中的小鼠胚胎和成年肾脏中表达的空间模式，2）进一步鉴定和表征相关的肾囊蛋白相互作用蛋白，3）确定肾囊蛋白对粘附调节的p130 Cas酪氨酸磷酸化和上皮细胞紧密连接形成和小管形成的影响。

The long-term objective of this proposal is to understand the nature of the cellular defects underlying the pathogenesis of familial juvenile nephronophthisis (NPH) -- a common genetic cause of kidney failure in children. Although clinical and histological observations indicate NPH results from a loss of normal excretory tubule function, the pathogenesis of the disease remains obscure. Recently a human gene mutated in the majority of NPH cases was isolated and the sequence of its encoded protein ("nephrocystin") was determined. A clue to the biochemical function of nephrocystin is the presence of a "Src homology 3" (SH3) domain known for mediating specific protein-protein interactions. New insight into the pathogenesis of NPH will now come from further study of nephrocystin. In preliminary studies, cDNAs encoding mouse nephrocystin have been isolated and the transcripts have been detected during post-implantation mouse embryogenesis and in a variety of adult tissues including the kidney. New information concerning the biochemical function of nephrocystin was obtained by identifying a tyrosine kinase substrate, pl30Cas, as a protein bound by the nephrocystin SH3 domain and by showing that nephrocystin localizes to lateral membranes of polarized epithelial cells. These observations led to our general hypothesis that nephrocystin functions in the morphogenesis and/or maintenance of the kidney tubular epithelium, requiring specific interactions with pl30Cas and other proteins involved in cell adhesive interactions. To test and expand the hypothesis, specific aims are proposed to: 1) determine the spatial pattern of expression of nephrocystin in developing mouse embryos and in the adult kidney, 2) further identify and characterize relevant nephrocystin-interacting proteins, and 3) determine the effects of nephrocystin on adhesion regulated p130Cas tyrosine phosphorylation and epithelial cell tight junction formation and tubulogenesis.