Angiotensin II‐induced stimulation of NHE3 is dependent on IRBIT and CaMKII

Abstract

NHE3 plays an essential role in the reabsorption of NaCl, HCO3−, and fluid in the proximal tubules of the kidney. Accumulating evidence has suggested that angiotensin II (ANG II) at physiological concentrations exerts a stimulatory effect on renal reabsorption of NaCl and filtered water by targeting NHE3. However, the underlying mechanisms remain incompletely elucidated. Herein, we aimed to determine the role of IRBIT in ANG II‐induced stimulation of NHE3 and to elucidate the underlying mechanisms. In OKP cells, 1 nM ANG II stimulated NHE3 activity by ~22%. In comparison, knockdown of IRBIT resulted in ~7% increase, whereas ectopic expression of IRBIT enhanced NHE3 activity by ~33%. Furthermore, ANG II induced over 20% increase in the surface NHE3 expression in OKP cells as determined by surface biotinylation, which was confirmed by immunofluorescence labeling of NHE3 on the apical membrane. CaMKII was shown to phosphorylate IRBIT and, hence, we tested whether CaMKII is required for ANG II‐activation of NHE3. Our results showed that inhibition of CaMKII by KN‐93 or ectopic expression of the natural inhibitor protein of CaMKII resulted in significant suppression of NHE3 activation by ANG II. In conclusion, our current results demonstrate that ANG II regulates NHE3 via CaMKII‐dependent mechanism and identify IRBIT as a physiological regulator of NHE3 in renal proximal epithelial cells.