Abstract
Two types of smooth muscle myosin heavy chain (MHC) isoforms, SM1 and SM2, were recently identified to have different carboxyl termini (Nagai, R., Kuro-o, M., Babij, P., and Periasamy, M. (1989) J. Biol. Chem. 264, 9734-9737). SM1 and SM2 are considered to be generated from a single gene through alternative RNA splicing. In this study we investigated expression of vascular MHC isoforms during development in rabbits at the mRNA, protein, and histological levels. In adults, all smooth muscle cells reacted with both anti-SM1 and anti-SM2 antibodies on immunofluorescence, suggesting the coexpression of SM1 and SM2 in a single cell. In fetal and perinatal rabbits, however, only anti-SM1 antibody consistently reacted with smooth muscles. Reactivity with anti-SM2 antibody was negative in the fetal and neonatal blood vessels and gradually increased during 30 days after birth. These developmental changes in SM1 and SM2 expression at the histological level coincided with mRNA expression of each MHC isoform as determined by S1 nuclease mapping, indicating that expression of SM1 and SM2 is controlled at the level of RNA splicing. However, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of myosin from fetal and perinatal aortas revealed the presence of large amount of SM2. Interestingly, fetal SM2 did not cross-react with our anti-SM2 antibody on immunoblotting. We conclude that expression of SM1 and SM2 are differentially regulated during development and that a third type of MHC isoform may exist in embryonic and perinatal vascular smooth muscles.
Highlights
From $the 3rd Department of Internal Medicine and the §Department of Laboratory Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo, Japan 113and the
AlternativeRNA Splicing.In this study we investigated In order better to understand smooth muscle myosin heavy chain (MHC) gene expression of vascular MHC isoforms during develop- expression, we tried inthis study to determine whetheror not ment in rabbits at themRNA, protein, and histological levels
ImmunologicalAnalysis of Smooth Muscle MHC Zsoforms-Previously we reported anti-SM1 and anti-SM2 antibodies, C1 and C2, raised in rats using short peptides which are specific to the carboxyl anti-SM2 antibodyonimmunoblotting
Summary
MHC heterogeneity has not been well defined until quite recently. Nagai et al [11,12] succeeded in isolating two types of smooth muscle MHC cDNA clones, SMHC40 and SMHC29, and suggested that SMHC40 and SMHC29 mRNAs originate from a single smooth muscle. (MHC) isoforms, SM1 and SM2, were recently identi- raised antibodies againsttwo synthetic peptides deduced from fied tohavedifferentcarboxyltermini Expression of smooth muscle MHC isoforms is subject to developmental regulation as known for the cardiac and the skeletal muscle MHCs. Our results clearly demonstrate that expression of SM1 and SM2 is developmentally regulated in only anti-SM1 antibody consistently reactedthe vascular system. Reactivity with anti-SM2 anti- existence of a third type MHC isoform in the embryonic and body was negative in the fetal andneonatalblood perinatal vascular smooth muscles.
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