pH-dependent regulation of the α-subunit of H⁺-K⁺-ATPase (HKα₂)

Abstract

: 1745-1755, 2010). In the present study, we investigated the ability of chronic acidosis and GPR4 to regulate HK␣2 expression in HEK-293 cells. Chronic acidosis was modeled in vitro by using multiple methods: reducing media pH by adjusting bicarbonate concentration, adding HCl, or by increasing the ambient concentration of CO 2. PKA activity and HK␣2 protein were monitored by immunoblot analysis, and HK␣ 2 mRNA, by real-time PCR. Chronic acidosis did not alter the expression of HK␣ 2 mRNA; however, PKA activity and HK␣ 2 protein abundance increased when media pH decreased from 7.4 to 6.8. Furthermore, this increase was independent of the method used to create chronic acidosis. Heterologous expression of GPR4 was sufficient to increase both basal and acid-stimulated PKA activity and similarly increase basal and acidstimulated HK␣ 2 expression. Collectively, these results suggest that chronic acidosis and GPR4 increase HK␣ 2 protein by increasing PKA activity without altering HK␣ 2 mRNA abundance, implicating a regulatory role of pH-activated GPR4 in homeostatic regulation of HK␣ 2 and acid-base balance.

Adaptation by the collecting duct to an exogenous acid load is blunted by deletion of the proton-sensing receptor GPR4

“…We did not observe such differences in the response of the apical transporters pendrin and H ϩ -ATPase. It seems unlikely to anticipate identification of a single or unique acid sensor in the kidney because current evidence suggests a complex network of kinases [e.g., Pyk2 adhesion kinase (34,39,58), ErbB1/2 tyrosine kinase (87), and AMPK (2)], enzymes [e.g., ammoniagenic enzymes (24,25,43) and soluble adenylyl cyclase (14,64)], G protein-coupled receptors like GPR4 (23,88) or OGR1 (63) or CaSR (28,73), the insulin receptor-related receptor (InsR-RR) (68), ion channels [e.g., TASK2 (101)], and transporters [e.g., vATPase (2,62)], which may all be involved in the response to a reduction in pH. Furthermore, multiple sensors may offer advantages based on different ranges of pH sensitivity, or cellular and subcellular localization.…”

“…Protonsensing receptors (GPR4, OGR1, TDAG8) constitute a small, unique family of G protein-coupled receptors that accept protons as ligands and are activated by protons in a physiological pH range to induce production of cAMP (GPR4 and TDAG8) or inositol 1,4,5-trisphosphate (IP 3 ; OGR1) in a pH-dependent manner (51,59). Recently this unique family of GPCRs has been linked to pH sensor function in a number of tissues and organs, including bone (36,37), developing blood vessels and endothelial cells (106), airway smooth muscle cells (79), kidney (23,88), and to pathological conditions such as cancer (17,26,53,86), ischemia, inflammation (20,27), and acid-base disturbances (23,63,88). Of the three receptors, only GPR4 is relatively abundant in the kidney.…”

Emerging roles for the pH-sensing G protein-coupled receptors in response to acidotic stress

“…59 In renal HEK293 epithelial cells GPR4 overexpression was found to increase the activity of PKA. 60 In addition, the protein expression of H + -K + -ATPase α-subunit (HKα 2 ) was increased following GPR4 overexpression dependent on increased PKA activity. 60 GPR68 has also been reported to alter proton export of HEK293 cells by stimulating the Na + /H + exchanger and H + -ATPase.…”

“…60 In addition, the protein expression of H + -K + -ATPase α-subunit (HKα 2 ) was increased following GPR4 overexpression dependent on increased PKA activity. 60 GPR68 has also been reported to alter proton export of HEK293 cells by stimulating the Na + /H + exchanger and H + -ATPase. 58 The activation of GPR68 by acidosis was found to stimulate this effect through a cluster of extracellular histidine residues and the G q /PKC signaling pathway.…”

“…[50][51][52] Renal N/A Regulates renal acid excretion in mice and H + -K + -ATPase expression. 59,60 Regulates proton transporter activity and proton extrusion in the kidney. 58 …”