CArG elements control smooth muscle subtype–specific expression of smooth muscle myosin in vivo
Ichiro Manabe, Gary K. Owens
Ichiro Manabe, Gary K. Owens
Published April 1, 2001
Citation Information: J Clin Invest. 2001;107(7):823-834. https://doi.org/10.1172/JCI11385.
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Article

CArG elements control smooth muscle subtype–specific expression of smooth muscle myosin in vivo

Abstract

Expression of smooth muscle myosin heavy chain (SM-MHC) is tightly controlled depending on the differentiated state of smooth muscle cells (SMCs). To better understand the mechanisms that regulate transcription of the SM-MHC gene in vivo, we tested the function of several conserved CArG elements contained within the –4200 to +11600 region of this gene that we had previously shown to drive SMC-specific expression in transgenic mice. CArG1 in the 5′-flanking sequence was required for all SMCs, while CArG2 and a novel intronic CArG element were differentially required in SMC subtypes. Of particular note, mutation of the intronic CArG selectively abolished expression in large arteries. A promoter construct containing three repeats of a conserved 227-bp intronic CArG-containing region was sufficient to direct transcription in vascular SMCs in transgenic mice, although this construct was also expressed in skeletal and cardiac muscle. These results support a model in which transcriptional regulation of SM-MHC is controlled by multiple positive and negative modular control regions that differ between SMCs and non-SMCs and among SMC subtypes. We also demonstrated that the CArG elements of the endogenous SM-MHC gene were bound by SRF in chromatin.

Authors

Ichiro Manabe, Gary K. Owens

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Figure 3

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Large artery-specific silencing of the reporter gene in intronic CArG-mu...
Large artery-specific silencing of the reporter gene in intronic CArG-mutant mice. Abdominal organs removed en block showed reporter expression in the blood vessels and urinary tract in the wild-type (a) and intronic CArG-mutant (b) transgenic mice. To better illustrate transgene expression in large arteries, several smaller arteries and connective tissues were removed, and the tissues were cleared. The supramesenteric artery, which was stained positive, was removed from the intronic CArG-mutant mouse tissues. A portion of the tissues is expanded in the inset in b. Arrowheads indicate the position of aorta that is not visible because of the lack of staining. Note that the blood vessels within the kidneys were not stained in either wild-type or intronic CArG mutants. (c and d) The thoracic aorta and branching arteries of the wild-type (c) and the intronic CArG-mutant (d) transgenic mice. (e) View of the large arteries in the cervicothoracic region of the intronic CArG-mutant transgenic mouse. (f) The large arteries and their branches in the abdomen of the intronic CArG mutant. A portion of the arteries is expanded in the inset in (f). (g–j) Histological examination of the abdominal aorta and inferior vena cava of the wild-type (g, i) and the intronic CArG mutant (h, j). The boxed areas (g, h) are shown by a higher magnification (i, j). Ao, aorta; DA, ductus arteriosus; IVC, inferior vena cava; SCA, subclavian artery.