A consensus sequence in the endothelin-B receptor second intracellular loop is required for NHE3 activation by endothelin-1

Endothelin-1 (ET-1) increases the activity of Na⁺/H⁺ exchanger 3 (NHE3), the major proximal tubule apical membrane Na ⁺/H⁺ antiporter. This effect is seen in opossum kidney (OKP) cells expressing the endothelin-B (ET_B) and not in cells expressing the endothelin-A (ETA) receptor. However, ET-1 causes similar patterns of protein tyrosine phosphorylation, adenylyl cyclase inhibition, and increases in cell [Ca²⁺] in ET_A- and ET _B-expressing OKP cells, implying that an additional mechanism is required for NHE3 stimulation by the ET_B receptor. The present studies used ET_A and ET_B receptor chimeras and site-directed mutagenesis to identify the ET receptor domains that mediate ET-1 regulation of NHE3 activity. We found that binding of ET-1 to the ET_A receptor inhibits NHE3 activity, an effect for which the COOH-terminal tail is necessary and sufficient. ET-1 stimulation of NHE3 activity requires the COOH-terminal tail and the second intracellular loop of the ET_B receptor. Within the second intracellular loop, a consensus sequence was identified, KXXXVPKXXXV, that is required for ET-1 stimulation of NHE3 activity. This sequence suggests binding of a homodimeric protein that mediates NHE3 stimulation.