title: Over-expressing transient receptor potential cation channel 6 in podocytes induces cytoskeleton rearrangement through increases of intracellular Ca2+ and RhoA activation.
abstract: Transient receptor potential cation channel 6 (TRPC6) is one of the key molecules for filtration barrier function of podocytes. Over-expression of TRPC6 in podocytes is frequently found in acquired or inherited proteinuric kidney diseases, and animal model over-expression of TRPC6 may lead to proteinuria. To investigate the impact of TRPC6 over-expression in podocytes on its function and its relation to proteinuria in kidney diseases, we over-expressed TRPC6 in mouse podocytes by transient transfection of TRPC6 cDNA plasmid, and observed their changes in foot processes, intracellular F-actin distribution, nephrin and synaptopodin expression, electrophysiology, RhoA activity and intracellular Ca(2+). In podocytes over-expressing TRPC6, cell processes were reduced remarkably in association with the derangement of cytoskeleton demonstrated by the abnormal distribution of intracellular F-actin. These cells also displayed a higher increase of intracellular Ca(2+) ion to the TRPC6 agonist 1-oleoyl-acetyl-sn-glycerol and a higher current in the patch-clamp experiment, down-regulation of nephrin and synaptopodin expression and increase of activated RhoA. These changes could be rescued by the treatment of the cells with U73122 to block TRPC6 channel or BAPTA-AM to chelate intracellular Ca(2+) ion. Additionally, the podocytes over-expressing TRPC6 treated with RhoA inhibitor Y-27632 showed an improvement in F-actin arrangement in the cells and increase of nephrin and synaptopodin expression. From these results, we therefore propose that over-expression of TRPC6 in podocytes may be one of the fundamental changes relating to the dysfunction of the slit diaphragm and proteinuria. Podocytes over-expressing TRPC6 may lead to higher intracellular Ca(2+) ion concentration in the presence of stimuli. The increase of intracellular Ca(2+) down-regulates the expression of two important molecules, nephrin on slit diaphragm and synaptopodin in cytoskeleton, and stimulates RhoA activity, which in turn causes F-actin derangement and the decrease of foot processes.
authors.affiliation: Department of Pediatrics, Peking University First Hospital, Beijing 100034, China.
authors.forename: Lina
authors.initials: L
authors.lastname: Jiang
authors.forename: Jie
authors.initials: J
authors.lastname: Ding
authors.forename: Haojan
authors.initials: H
authors.lastname: Tsai
authors.forename: Lin
authors.initials: L
authors.lastname: Li
authors.forename: Quancheng
authors.initials: Q
authors.lastname: Feng
authors.forename: Jing
authors.initials: J
authors.lastname: Miao
authors.forename: Qingfeng
authors.initials: Q
authors.lastname: Fan
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002413:Cations, Divalent / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism*; D015870:Gene Expression; D008565:Membrane Proteins / Q000096:biosynthesis; D051379:Mice; D008840:Microfilament Proteins / Q000096:biosynthesis; D050199:Podocytes / Q000378:metabolism*; D050052:TRPC Cation Channels / Q000096:biosynthesis*; D000074287:TRPC6 Cation Channel; D020741:rho GTP-Binding Proteins / Q000378:metabolism*; D020742:rhoA GTP-Binding Protein
chemical_list: D002413:Cations, Divalent; D008565:Membrane Proteins; D008840:Microfilament Proteins; C503968:Synpo protein, mouse; D050052:TRPC Cation Channels; D000074287:TRPC6 Cation Channel; C492146:Trpc6 protein, mouse; C113267:nephrin; C526380:RhoA protein, mouse; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein; D002118:Calcium