Kotsis Fruzsina, Nitschke Roland, Boehlke Christopher, Bashkurov Mikhail, Walz Gerd, Kuehn E Wolfgang
Department of Medicine, University Hospital Freiburg, Renal Unit, Hugstetter Str. 55, Freiburg, Germany.
Pflugers Arch. 2007 Mar;453(6):819-29. doi: 10.1007/s00424-006-0168-0. Epub 2007 Jan 5.
Primary cilia have been shown to play an important role in embryonic development as well as in postnatal life. Dysfunctional cilia are associated with situs inversus, retinal abnormalities, impaired mucociliary clearance, infertility, hydrocephalus, and congenital renal cysts. In autosomal dominant polycystic kidney disease, mutations of the ciliary proteins polycystin1 or the transient receptor potential (TRP) channel family protein polycystin2 (TRPP2) cause progressive cyst formation and destruction of the kidney. Primary cilia act as flow sensors and respond to flow-mediated bending with a prolonged intracellular calcium increase, which appears to require an intact polycystin protein complex. We have established a novel flow chamber system, which allows us to study renal epithelial cells by live cell imaging. We show that MDCK cells respond to flow by a delayed increase in intracellular calcium and that this response requires these cells to be ciliated. We show that a novel interactor of TRPP2, kidney injury molecule-1 (Kim1), which is expressed at low levels in the normal kidney and upregulated after ischemia, in renal cell cancer and in PKD is targeted to primary cilia when stably expressed in MDCK cells. We demonstrate that expression of tyrosine mutant Kim1, lacking a conserved tyrosine in the intracellular tail, abolishes the calcium increase in response to flow in a dominant negative manner. These results establish Kim1 as a novel regulatory molecule of flow-induced calcium signaling.
原发性纤毛已被证明在胚胎发育以及出生后生活中发挥重要作用。功能失调的纤毛与内脏反位、视网膜异常、黏液纤毛清除功能受损、不孕、脑积水和先天性肾囊肿有关。在常染色体显性多囊肾病中，纤毛蛋白多囊蛋白1或瞬时受体电位（TRP）通道家族蛋白多囊蛋白2（TRPP2）的突变会导致肾脏逐渐形成囊肿并遭到破坏。原发性纤毛充当流量传感器，并对流量介导的弯曲作出反应，使细胞内钙持续增加，这似乎需要完整的多囊蛋白复合物。我们建立了一种新型流动腔系统，使我们能够通过活细胞成像研究肾上皮细胞。我们发现，MDCK细胞通过细胞内钙的延迟增加对流量作出反应，并且这种反应要求这些细胞有纤毛。我们发现，TRPP2的一种新型相互作用分子——肾损伤分子1（Kim1），在正常肾脏中低水平表达，在缺血后、肾细胞癌和多囊肾病中上调，当在MDCK细胞中稳定表达时靶向原发性纤毛。我们证明，在细胞内尾部缺乏保守酪氨酸的酪氨酸突变型Kim1的表达，以显性负性方式消除了对流量作出反应的钙增加。这些结果确立了Kim1作为流量诱导钙信号传导的一种新型调节分子的地位。
