Role of protein kinase C in desensitization of rat endothelin A receptors

Rat anterior pituitary cells express the calcium‐mobilizing endothelin‐A (ETA) receptors, which activation is accompanied with a rapid desensitization and internalization through still not well‐defined pathways. Here we examined the role of protein kinase C (PKC) signaling pathway in activation and desensitization of the rat ETA receptors when expressed in HEK293 cells. The coupling of these receptors to the calcium‐mobilizing pathway was almost abolished in cells pretreated with phorbol ester PMA, a PKC activator, whereas the desensitization of receptors was dramatically attenuated in cells pretreated with Ro31‐8425, a PKC specific inhibitor. To address the hypothesis that PKC‐mediated phosphorylation of receptors accounts for these effects, the four putative intracellular PKC phosphorylation sites at S289, S373, T416 and S420, which are conserved in all mammalian ETA receptors, were mutated to alanine. Mutations did not affect the plasma membrane expression and the calcium‐mobilizing action of receptors. However, the S373A mutant did not desensitize during the repetitive agonist application and phorbol ester/Ro31‐8425 treatments were ineffective, in contrast to the S289A, T416A, and S420A mutant receptors. These results suggest that the ETA receptor mediated Gq activation is desensitized via S373 phosphorylation mediated by PKC. Supported by the Intramural Research Program of the NICHD, NIH.