Cloning and Characterization of the Mouse α1C/A-Adrenergic Receptor Gene and Analysis of an α1C Promoter in Cardiac Myocytes: Role of an MCAT Element That Binds Transcriptional Enhancer Factor-1 (TEF-1)
- Cardiology Division and Research Service, Veterans Affairs Medical Center, San Francisco, California; and the Cardiovascular Research Institute and Department of Medicine, University of California, San Francisco, California
Abstract
α1-Adrenergic receptor (AR) subtypes in the heart are expressed by myocytes but not by fibroblasts, a feature that distinguishes α1-ARs from β-ARs. Here we studied myocyte-specific expression of α1-ARs, focusing on the subtype α1C (also called α1A), a subtype implicated in cardiac hypertrophic signaling in rat models. We first cloned the mouse α1C-AR gene, which consisted of two exons with an 18 kb intron, similar to the α1B-AR gene. The receptor coding sequence was >90% homologous to that of rat and human. α1C-AR transcription in mouse heart was initiated from a single Inr consensus sequence at −588 from the ATG; this and a putative polyadenylation sequence 8.5 kb 3′ could account for the predominant 11 kb α1C mRNA in mouse heart. A 5′-nontranscribed fragment of 4.4 kb was active as a promoter in cardiac myocytes but not in fibroblasts. Promoter activity in myocytes required a single muscle CAT (MCAT) element, and this MCAT bound in vitro to recombinant and endogenous transcriptional enhancer factor-1. Thus, α1C-AR transcription in cardiac myocytes shares MCAT dependence with other cardiac-specific genes, including the α- and β-myosin heavy chains, skeletal α-actin, and brain natriuretic peptide. However, the mouse α1C gene was not transcribed in the neonatal heart and was not activated by α1-AR and other hypertrophic agonists in rat myocytes, and thus differed from other MCAT-dependent genes and the rat α1C gene.
Footnotes
-
Send reprint requests to: Paul C. Simpson, M.D., VAMC 111-C-84150, Clement St., San Francisco, CA 94121. E-mail: pcs{at}itsa.ucsf.edu
-
↵1 Current address: Department of Medicine, University of Louisville, Louisville, Kentucky.
-
↵2 Throughout this article, we use the name α1C-AR to avoid potential ambiguities with the name α1A (Graham et al., 1996). For example, the cloned α1d-AR was named originally the α1A (Lomasney et al., 1991) and the accession numbers in GenBank for the α1D sequence (M60654 and M60655) still refer to this gene as the α1A.
-
This work was supported by a Fellowship from the Cardiovascular Research Institute, the University of California, San Francisco (T32HL07731) (T.D.O.); a Fellowship from the American Heart Association, Western States Affiliate (D.G.R.); and the Department of Veterans Affairs and the National Institutes of Health (P.C.S.). The new cloned sequences reported herein have been submitted to GenBank under accession numbers AF362076, AF362077, and AF362078.
- Abbreviations:
- AR
- adrenergic receptor
- TEF-1
- transcriptional enhancer factor 1
- PCR
- polymerase chain reaction
- BAC
- bacterial artificial chromosome
- bp
- base pair(s)
- kb
- kilobase pair(s)
- RPA
- ribonuclease protection assay
- PMA
- phorbol-12-myristate-13-acetate
- EC
- enhancer core
- PGF2α
- prostaglandin F2α
- CAT
- chloramphenicol acetyltransferase
- tk
- thymidine kinase
- CRE
- cAMP response element
-
- Received August 29, 2000.
- Accepted January 17, 2001.
- The American Society for Pharmacology and Experimental Therapeutics
