Specific Pregnancy-Induced Angiotensin II Type-1 Receptor Expression in Ovine Uterine Artery Does Not Involve Formation of Alternate Splice Variants or Alternate Promoter Usage¹

^c Departments of Obstetrics & Gynecology, Perinatal Research Laboratories, and ^d Meat & Animal Science, University of Wisconsin-Madison, Madison, Wisconsin 53715

Recently we reported that pregnancy is associated with a dramatic increase in angiotensin II type-1 receptor (AT₁-R; both protein and mRNA) in ovine uterine artery endothelial cells (UAEC), which far exceeds that seen in omental (systemic) arteries. Recent reports also suggest that alternate splicing of AT₁-R mRNA may play a role in regulation of AT₁-R expression in humans. Herein, we have investigated the possibility of alternate transcript splicing/promoter usage in UAEC from pregnant ewes by 5'-RACE (rapid amplification of cDNA 5'-ends). To provide our control "reference" sequences, we first performed 5'-RACE analysis of AT₁-R mRNA transcripts in liver, kidney, and adrenal cortex. Analysis of 17 resultant clones showed exceptional homology, indicating that a single identically spliced mRNA product is observed in all three ovine tissues. Homology of the 5'-untranslated region to that of the human was low (34.2%), but four in-context start/stop codons and the beginning of human exons 1 and 5 were highly conserved. Subsequently we isolated 30 individual clones using UAEC RNA from three pregnant ewes and found no evidence of any sequence formed through unique splicing or promoter usage. We conclude that the pregnancy-induced increase in AT₁-R expression unique to UAEC during pregnancy is not mediated by splicing of a unique transcript or unique promoter usage.

¹ These studies were supported by grants AHA-WI 95-GB-41, USDA 9601773, and NIH HL56702 to I.M.B. and HD33255, HL49210, and HL57653 to R.R.M.

² Correspondence: Ian M. Bird, University of Wisconsin-Madison, Department Obstetrics & Gynecology, Perinatal Research Laboratories, 7E Meriter Hospital/Park, 202 South Park St., Madison, WI 53715. FAX: (608) 257–1304; imbird{at}facstaff.wisc.edu

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