title: Novel signaling mechanisms of intracellular angiotensin II-induced NHE3 expression and activation in mouse proximal tubule cells.
abstract: Expression of a cytosolic cyan fluorescent fusion protein of angiotensin II (ECFP/ANG II) in proximal tubules increases blood pressure in rodents. To determine cellular signaling pathways responsible for this response, we expressed ECFP/ANG II in transport-competent mouse proximal convoluted tubule cells (mPCT) from wild-type (WT) and type 1a ANG II receptor-deficient (AT(1a)-KO) mice and measured its effects on intracellular ANG II levels, surrogates of Na/H exchanger 3 (NHE3)-dependent Na(+) absorption, as well as MAP kinases and NF-κB signaling. In WT mPCT cells, ECFP/ANG II expression doubled ANG II levels, increased NHE3 expression and membrane phospho-NHE3 proteins threefold and intracellular Na(+) concentration by 65%. These responses were associated with threefold increases in phospho-ERK 1/2 and phospho-p38 MAPK, fivefold increases in p65 subunit of NF-κB, and threefold increases in phospho-IKKα/β (Ser 176/180) proteins. These signaling responses to ECFP/ANG II were inhibited by losartan (AT(1) blocker), PD123319 (AT(2) blocker), U0126 (MEK1/MEK2 inhibitor), and RO 106-9920 (NF-κB inhibitor). In mPCT cells of AT(1a)-KO mice, ECFP/ANG II also increased the levels of NHE3, p-ERK1/2, and p65 proteins above their controls, but considerably less so than in WT cells. In WT mice, selective expression of ECFP/ANG II in vivo in proximal tubules significantly increased blood pressure and indices of sodium reabsorption, in particular levels of phosphorylated NHE3 protein in the membrane fraction and proton gradient-stimulated (22)Na(+) uptake by proximal tubules. We conclude that intracellular ANG II may induce NHE3 expression and activation in mPCTs via AT(1a)- and AT(2) receptor-mediated activation of MAP kinases ERK 1/2 and NF-κB signaling pathways.
authors.affiliation: Laboratory of Receptor and Signal Transduction, Department of Pharmacology and Toxicology, University of Mississippi Medical Center, Jackson, Mississippi 39216-4505, USA.
authors.forename: X C
authors.initials: XC
authors.lastname: Li
authors.forename: U
authors.initials: U
authors.lastname: Hopfer
authors.forename: J L
authors.initials: JL
authors.lastname: Zhuo
mesh_terms: D000804:Angiotensin II / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D049452:Green Fluorescent Proteins / Q000378:metabolism; D066298:In Vitro Techniques; D007687:Kidney Tubules, Proximal / Q000166:cytology* / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000502:physiology; D051379:Mice; D018345:Mice, Knockout; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D044140:Receptor, Angiotensin, Type 1 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D044139:Receptor, Angiotensin, Type 2 / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
chemical_list: C485182:Cyan Fluorescent Protein; D016328:NF-kappa B; D044140:Receptor, Angiotensin, Type 1; D044139:Receptor, Angiotensin, Type 2; C000613386:Slc9a3 protein, mouse; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; D000804:Angiotensin II; D049452:Green Fluorescent Proteins; D020929:Mitogen-Activated Protein Kinase Kinases