Cloning and functional identification of a novel rat endothelin receptor type A isoform

Abstract

Endothelins (ET) act via two receptors, ETAR and ETBR, to regulate variable cellular functions in numerous tissues. In anterior pituitary, the BQ123-sensitive ETAR are present in all secretory cell types and are involved in the control of calcium signaling and hormone secretion. However, in gonadotrophs but not in lactotrophs these receptors rapidly desensitize, raising the possibility of the existence of pituitary cell type-specific ETAR variants. Here, we report about the cloning and expression of a novel ETAR isoform from rat anterior pituitary, termed ETAR-C13. The cloned cDNA has an inserted fragment between the exons 6 and 7 of wild type ETAR, resulting in substitution of C-terminal 382S-426N sequence of wild type receptor with shorter 382A-399L sequence of spliced receptor. The putative domain of C-terminal sequence responsible for coupling to Gq/11 proteins was altered, and the predicted protein kinase C and casein kinase 2 phosphorylation sites were deleted in ETAR-C13. The mRNA transcripts for ETAR-C13 were identified in immortalized and cultured pituitary cells and pituitary tissue, but not in other tissues. The 125I-ET-1 binding at ETAR-C13 expressed in HEK293 cells was completely displaced by BQ123, the coupling to Gq/11 proteins was preserved but was shifted rightward, and the coupling to Gs proteins was significantly attenuated. Furthermore, the rate of ETAR-C13 desensitization and internalization was lower compared to wild type ETAR. These results indicate that the expression of ETAR-C13 in lactotrophs and somatotrophs probably accounts for the long-lasting signaling. Supported by the Intramural Research Program of the NICHD, NIH.