Microarray Profiling of G Protein‐coupled Receptor Signaling Responses to Overexpression of An Intracellular Angiotensin II Fusion Protein in Mouse Proximal Tubule Cells

Abstract

We have recently demonstrated that overexpression of an intracellular angiotensin II (ANG II) fusion protein, ECFP/ANG II, selectively in the proximal tubule of the kidney increases blood pressure in rats and mice, but specific signaling mechanisms involved remain unknown. The present study used the Mouse GPCR Signaling Pathway Microarray to determine GPCR signaling responses to overexpression of ECFP/ANG II in mouse proximal tubule cells (mPCT). mPCTs from wild‐type and AT1a‐KO mice were transfected with or without ECFP/ANG II for 48 h, and RNA samples were extracted for microarray analysis of GPCR signaling responses to ECFP/ANG II (n=6 per group). In wild‐type mPCT cells, overexpression of ECFP/ANG II significantly upregulated dozens of GPCR signaling proteins (p<0.05). Significantly increased GPCR expression or signaling proteins included Agtrl1, Agtr2, Agtrap, Arrb1, Arrb2, Calcr, Casr, Ccl2, Ccl4, Cmklr1, Drd1a, Drd2, Edg6, Fgf2, IL4, NOS2, Nppa, Pthr1, and Vegfa etc. By contrast, significantly decreased GPCR signaling proteins included Adcy5, Calcr1, Cdkn1a, Ctgf, Cyp19a1, Edn1, Gcgr, Il1r1, Pik3cg, and Ucp1 etc. The effects of ECFP/ANG II overexpression in wild‐type mPTCs were significantly attenuated in mPCT cells of AT1a‐KO mice, which included Agtrl1, Arrb1, Arrb2, Bcl2l1, Casr, Ccl2, Ccl4, Drd2, Nppa, Pthr1, and Vegfa. Other GPCRs or signaling proteins were not significantly altered in mPCT cells of AT1a‐KO mice. These results suggest that the GPCR Signaling Pathway Microarray may be a useful approach to identify both GPCR‐dependent and GPCR‐independent signaling mechanisms underlying intracellular ANG II‐induced responses in proximal tubule cells.