title: Human kidney-2 cells harbor functional dopamine D1 receptors that require Giα for Gq/11α signaling.
abstract: A recent study demonstrated that the dopamine D1 receptor (D1R) is nonfunctional in human kidney cells, HK2 cells, in terms of their inability to couple to Gs protein in response to the D1R agonist fenoldopam. Since D1R also couples to Gq protein, we tested whether D1R is functional in HK2 cells in terms of their ability to couple to Gq and produce downstream signaling. For comparison, we also studied another receptor, angiotensin II type 1 receptor (AT1R) known to couple to Gq. Protein kinase C (PKC) and (86)rubidium transport activities were determined as surrogate downstream signaling markers. Fenoldopam and angiotensin II increased PKC activity, which decreased in the presence of respective receptor antagonists (SCH23390 for D1R; candesartan for AT1R), PKC (chelerythrine chloride) and Gi protein (pertussis toxin) inhibitors and Gq/11α siRNA. Furthermore, fenoldopam and angiotensin II increased (35)S-GTPγS binding, an index of receptor-G protein coupling, which decreased with pertussis toxin and in Gq/11α-depleted cells. Also, fenoldopam-mediated inhibition of (86)rubidium transport (an index of Na-K-ATPase activity) was attenuated with SCH23390, chelerythrine chloride, pertussis toxin, and Gq/11α siRNA. Moreover, fenoldopam caused a decrease in cytosolic and increase in membranous abundance of Gq/11α. The immunoprecipitated levels of Gq/11α in the membranes were greater in fenoldopam-treated cells, and Giα coimmunoprecipitated with Gq/11α. Our results suggest that both D1R and AT1R are functional in HK2 cells, enabling Gq-mediated downstream signaling in a Gi dependent manner.
authors.affiliation: Heart and Kidney Institute, College of Pharmacy, Univ. of Houston, Houston, Texas 77204. masghar@uh.edu.
authors.forename: Indira D
authors.initials: ID
authors.lastname: Pokkunuri
authors.forename: Gaurav
authors.initials: G
authors.lastname: Chugh
authors.forename: Mohammad
authors.initials: M
authors.lastname: Asghar
mesh_terms: D001692:Biological Transport; D018929:Cell Culture Techniques; D004847:Epithelial Cells / Q000378:metabolism; D018818:Fenoldopam / Q000494:pharmacology; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D006801:Humans; D007668:Kidney / Q000187:drug effects / Q000378:metabolism*; D007687:Kidney Tubules, Proximal / Q000187:drug effects / Q000378:metabolism*; D044140:Receptor, Angiotensin, Type 1 / Q000378:metabolism; D017447:Receptors, Dopamine D1 / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D044140:Receptor, Angiotensin, Type 1; D017447:Receptors, Dopamine D1; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D018818:Fenoldopam
keywords: D1R; Giα; Gq/11α; HK2; kidney