Cloning and functional identification of novel endothelin receptor type A isoforms in pituitary.
Mammalian endothelin (ET) receptors, termed ET(A)R and ET(B)R, are derived from two intron-containing genes and the functional splice variants of ET(B)R but not ET(A)R have been identified. Here, we report about the isolation of cDNAs of ET(A)R transcripts from rat anterior pituitary, which are generated by alternative RNA splicing. Deletion of exon 2 and insertion of fragments from intron 1 and 2 accounted for formation of three misplaced proteins, whereas the insertion of a fragment from intron 6 resulted in generation of a functional plasma membrane receptor, termed ET(A)R-C13. In this splice variant, the C-terminal 382S-426N sequence of ET(A)R was substituted with a shorter 382A-399L sequence, resulting in alteration of the putative domains responsible for coupling to G(q/11) and G(s) proteins and the endocytotic recycling, as well as in deletion of the predicted protein kinase C/casein kinase 2 phosphorylation sites. The mRNA transcripts for ET(A)R-C13 were identified in normal and immortalized pituitary cells and several other tissues. The pharmacological profiles of recombinant ET(A)R and ET(A)R-C13 were highly comparable, but the coupling of ET(A)R-C13 to the calcium-mobilizing signaling pathway was attenuated, causing a rightward shift in the potency for agonist. Furthermore, the efficacy of ET(A)R-C13 to stimulate adenylyl cyclase signaling pathway and to internalize was significantly reduced. These results indicate for the first time the presence of a novel ET(A) splice receptor, which could contribute to the functional heterogeneity among secretory pituitary cell types.
Hatae
Noriyuki
N
Section on Cellular Signaling, National Institute of Child Health and Human Development (NICHD), National Institutes of Health (NIH), Building 49, Room 6A-36, 49 Convent Drive, Bethesda, Maryland 20892-4510, USA.
Aksentijevich
Nadia
N
Zemkova
Hana W
HW
Kretschmannova
Karla
K
Tomic
Melanija
M
Stojilkovic
Stanko S
SS
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D002478:Cells, Cultured; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D014644:Genetic Variation; D006801:Humans; D008969:Molecular Sequence Data; D010903:Pituitary Gland, Anterior / Q000502:physiology*; D020033:Protein Isoforms / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D044022:Receptor, Endothelin A / Q000235:genetics*; D044023:Receptor, Endothelin B / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic
D017931:DNA Primers; D018076:DNA, Complementary; D020033:Protein Isoforms; D044022:Receptor, Endothelin A; D044023:Receptor, Endothelin B