Jak2 and Ca 2+ /Calmodulin are Key Intermediates for Bradykinin B 2 Receptor-Mediated Activation of Na + /H + Exchange in KNRK and CHO Cells

Abstract

Na⁺/H⁺ exchangers are ubiquitous in mammalian cells, carrying out key functions, such as cell volume defense, acid-base homeostasis, and regulation of the cytoskeleton. We used two screening technologies(FLIPR and microphysiometry) to characterize the signal transduction pathway used by the bradykinin B₂ receptor to activate Na⁺/H⁺ exchange in two cell lines, KNRK and CHO. In both cell types, B₂ receptor activation resulted in rapid increases in the rate of proton extrusion that were sodium-dependent and could be blocked by the Na⁺/H⁺ exchange inhibitors EIPA and MIA or by replacing extracellular sodium with TMA. Activation of Na⁺/H⁺ exchange by bradykinin was concentration-dependent and could be blocked by the selective B₂ receptor antagonist HOE140, but not by the B₁ receptor antagonist des-Arg10-HOE140. Inhibitors of Jak2 tyrosine kinase (genistein and AG490) and of CAM (W-7 and calmidazolium) attenuated bradykinin-induced activation of Na⁺/H⁺ exchange. Bradykinin induced formation of a complex between CAM and Jak2, supporting a regulatory role for Jak2 and CAM in the activation of Na⁺/H⁺ exchange in KNRK and CHO cells. We propose that this pathway (B₂ receptor → Jak2 → CAM → Na⁺/H⁺ exchanger) is a fundamental regulator of Na⁺/H⁺ exchange activity.