Isolation of a Full Length Ovine Angiotensin II Type-1 Receptor (AT1-R) cDNA

Abstract

A full-length ovine AT1-R cDNA (2358 bases: Genebank AF254119) was isolated from ovine adrenal cortex using 3′-RACE. While homology of the 5′-untranslated region to human sequence was low (34.2%), the beginning of both human exon 1 and human exon 5 (encoding the end of the 5′-untranslated sequence/complete protein coding region/3′-untranslated sequence) were exceptionally conserved and in context. The intervening untranslated sequence showed lower homology to human sequence, but still contained four additional ATG sequences close to corresponding “in frame” TGA Stop codons shown in human to impair AT1-R translation in vitro. The putative protein coding sequence was >99% identical to the previous reported ovine genomic sequence. The predicted amino acid sequence in turn encoded a protein with the properties of a seven α-helix transmembrane receptor sharing closest homology (99.1 %) to the bovine receptor and lowest to the rat Type 1a (90.5%). The 3′-untranslated region showed relatively high homology to the porcine and bovine receptor cDNA, but did not share the additional 643 bases found only in the bovine 3′-untranslated region. The ovine 3′-sequence included a polyadenylation signal as well as three AUUUA destabilization sequences observed in most other species including human. Thus ovine AT1-R mRNA stability may be short lived, and control of degradation may be an additional mechanism for regulation of ovine AT1-R expression. Supported by AHA-WI 95-GB-41, 97 GB 95 and USDA 9601773