Vasopressin receptor-mediated functional signaling pathway in primary cilia of renal epithelial cells

作者：Malay K. Raychowdhury, Arnolt J. Ramos, Peng Zhang, Margaret McLaughin, Xiao‐Qing Dai, Xing‐Zhen Chen, Nicolás Montalbetti, María del Rocío Cantero, Dennis A. Ausiello, Horacio F. Cantiello · 发表于：American Journal of Physiology-Renal Physiology · 年份：2008 · DOI：10.1152/ajprenal.90509.2008 · 被引用次数：71 · 研究领域：Genetic and Kidney Cyst Diseases、Biomedical Research and Pathophysiology、Organ Donation and Transplantation

The primary cilium of renal epithelial cells is a nonmotile sensory organelle, implicated in mechanosensory transduction signals. Recent studies from our laboratory indicate that renal epithelial primary cilia display abundant channel activity; however, the presence and functional role of specific membrane receptors in this organelle are heretofore unknown. Here, we determined a functional signaling pathway associated with the type 2 vasopressin receptor (V2R) in primary cilia of renal epithelial cells. Besides their normal localization on basolateral membrane, V2R was expressed in primary cilia of LLC-PK(1) renal epithelial cells. The presence of V2R in primary cilia was determined by spontaneous fluorescence of a V2R-gfp chimera and confirmed by immunocytochemical analysis of wild-type LLC-PK(1) cells stained with anti-V2R antibodies and in LLC-PK(1) cells overexpressing the V2R-Flag, with anti-Flag antibody. Ciliary V2R colocalized with adenylyl cyclase (AC) type V/VI in all cell types tested. Functional coupling of the receptors with AC was confirmed by measurement of cAMP production in isolated cilia and by testing AVP-induced cation-selective channel activity either in reconstituted lipid bilayers or subjected to membrane-attached patch clamping. Addition of either 10 microM AVP (trans) or forskolin (cis) in the presence but not the absence of ATP (1 mM, cis) stimulated cation-selective channel activity in ciliary membranes. This channel activity was reduced by addition...