© 1999 American Heart Association, Inc.
Rapid Communication |
Regulation of Smooth Muscle 
-Actin Expression In Vivo Is Dependent on CArG Elements Within the 5' and First Intron Promoter Regions
From the Department of Molecular Physiology and Biological Physics, University of Virginia Medical School, Charlottesville, Va.
Correspondence to Gary K. Owens, PhD, Box 449 Health Sciences Center, University of Virginia, Charlottesville, VA 22908.
Abstract—The aims of the
present studies were to define sufficient promoter sequences
required to drive endogenous expression of smooth muscle
(SM) 
-actin and to determine whether regulation of SM -actin
expression in vivo is dependent on CArG (CC(A/T)6GG)
cis elements. Promoter deletions and site directed
mutagenesis techniques were used to study gene regulation in transgenic
mice as well as in smooth muscle cell (SMC) cultures. Results
demonstrated that a Lac Z transgene that contained 547 bp of the 5' rat
SM -actin promoter was sufficient to drive embryonic expression of
SM -actin in the heart and in skeletal muscle but not in SMCs.
Transient transfections into SMC cultures demonstrated that the
conserved CArG element in the first intron had significant positive
activity, and gel shift analyses demonstrated that the intronic
CArG bound serum response factor. A transgene construct from -2600
through the first intron (p2600Int/Lac Z) was expressed in embryos and
adults in a pattern that closely mimicked endogenous SM
-actin expression. Expression in adult mice was completely
restricted to SMCs and was detected in esophagus, stomach, intestine,
lung, and nearly all blood vessels, including coronary,
mesenteric, and renal vascular beds. Mutation of CArG B completely
inhibited expression in all cell types, whereas mutation of the
intronic CArG selectively abolished expression in SMCs, which suggests
that it may act as an SMC-specific enhancer-like element. Taken
together, these results provide the first in vivo evidence for the
importance of multiple CArG cis elements in the
regulation of SM -actin expression.
Key Words: muscle, smooth • transgenic mice • transfection • gene expression
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C. P. Regan, I. Manabe, and G. K. Owens Development of a Smooth Muscle-Targeted Cre Recombinase Mouse Reveals Novel Insights Regarding Smooth Muscle Myosin Heavy Chain Promoter Regulation Circ. Res., September 1, 2000; 87(5): 363 - 369. [Abstract] [Full Text] [PDF]
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J. M. Miano, M. J. Carlson, J. A. Spencer, and R. P. Misra Serum Response Factor-dependent Regulation of the Smooth Muscle Calponin Gene J. Biol. Chem., March 24, 2000; 275(13): 9814 - 9822. [Abstract] [Full Text] [PDF]
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C. P. Mack, M. M. Thompson, S. Lawrenz-Smith, and G. K. Owens Smooth Muscle {alpha}-Actin CArG Elements Coordinate Formation of a Smooth Muscle Cell-Selective, Serum Response Factor-Containing Activation Complex Circ. Res., February 4, 2000; 86(2): 221 - 232. [Abstract] [Full Text] [PDF]
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F. Jung, A. D. Johnson, M. S. Kumar, B. Wei, M. Hautmann, G. K. Owens, and C. McNamara Characterization of an E-box-Dependent cis Element in the Smooth Muscle {alpha}-Actin Promoter Arterioscler Thromb Vasc Biol, November 1, 1999; 19(11): 2591 - 2599. [Abstract] [Full Text] [PDF]
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A. D. Johnson and G. K. Owens Differential activation of the SMalpha A promoter in smooth vs. skeletal muscle cells by bHLH factors Am J Physiol Cell Physiol, June 1, 1999; 276(6): C1420 - C1431. [Abstract] [Full Text] [PDF]
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B. Camoretti-Mercado, H.-W. Liu, A. J. Halayko, S. M. Forsythe, J. W. Kyle, B. Li, Y. Fu, J. McConville, P. Kogut, J. E. Vieira, et al. Physiological Control of Smooth Muscle-specific Gene Expression through Regulated Nuclear Translocation of Serum Response Factor J. Biol. Chem., September 22, 2000; 275(39): 30387 - 30393. [Abstract] [Full Text] [PDF]
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C. P. Mack, A. V. Somlyo, M. Hautmann, A. P. Somlyo, and G. K. Owens Smooth Muscle Differentiation Marker Gene Expression Is Regulated by RhoA-mediated Actin Polymerization J. Biol. Chem., January 5, 2001; 276(1): 341 - 347. [Abstract] [Full Text] [PDF]
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A. M. Hoggatt, A. M. Kriegel, A. F. Smith, and B. P. Herring Hepatocyte Nuclear Factor-3 Homologue 1 (HFH-1) Represses Transcription of Smooth Muscle-specific Genes J. Biol. Chem., September 29, 2000; 275(40): 31162 - 31170. [Abstract] [Full Text] [PDF]
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P. J. Adam, C. P. Regan, M. B. Hautmann, and G. K. Owens Positive- and Negative-acting Kruppel-like Transcription Factors Bind a Transforming Growth Factor beta Control Element Required for Expression of the Smooth Muscle Cell Differentiation Marker SM22alpha in Vivo J. Biol. Chem., November 22, 2000; 275(48): 37798 - 37806. [Abstract] [Full Text] [PDF]
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P. S. Chang, L. Li, J. McAnally, and E. N. Olson Muscle Specificity Encoded by Specific Serum Response Factor-binding Sites J. Biol. Chem., May 11, 2001; 276(20): 17206 - 17212. [Abstract] [Full Text] [PDF]
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B. P. Herring, A. M. Kriegel, and A. M. Hoggatt Identification of Barx2B, a Serum Response Factor-associated Homeodomain Protein J. Biol. Chem., April 20, 2001; 276(17): 14482 - 14489. [Abstract] [Full Text] [PDF]
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M. Strobeck, S. Kim, J. C. L. Zhang, C. Clendenin, K. L. Du, and M. S. Parmacek Binding of Serum Response Factor to CArG Box Sequences Is Necessary but Not Sufficient to Restrict Gene Expression to Arterial Smooth Muscle Cells J. Biol. Chem., May 4, 2001; 276(19): 16418 - 16424. [Abstract] [Full Text] [PDF]
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M. D'Addario, P. D. Arora, J. Fan, B. Ganss, R. P. Ellen, and C. A. G. McCulloch Cytoprotection against Mechanical Forces Delivered through beta 1 Integrins Requires Induction of Filamin A J. Biol. Chem., August 17, 2001; 276(34): 31969 - 31977. [Abstract] [Full Text] [PDF]
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C. J. Phiel, V. Gabbeta, L. M. Parsons, D. Rothblat, R. P. Harvey, and K. M. McHugh Differential Binding of an SRF/NK-2/MEF2 Transcription Factor Complex in Normal Versus Neoplastic Smooth Muscle Tissues J. Biol. Chem., September 7, 2001; 276(37): 34637 - 34650. [Abstract] [Full Text] [PDF]
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G. Schratt, U. Philippar, J. Berger, H. Schwarz, O. Heidenreich, and A. Nordheim Serum response factor is crucial for actin cytoskeletal organization and focal adhesion assembly in embryonic stem cells J. Cell Biol., February 18, 2002; 156(4): 737 - 750. [Abstract] [Full Text] [PDF]
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J. M. Miano, C. M. Kitchen, J. Chen, K. M. Maltby, L. A. Kelly, H. Weiler, R. Krahe, L. K. Ashworth, and E. Garcia Expression of human smooth muscle calponin in transgenic mice revealed with a bacterial artificial chromosome Am J Physiol Heart Circ Physiol, May 1, 2002; 282(5): H1793 - H1803. [Abstract] [Full Text] [PDF]
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I. Manabe and G. K. Owens Recruitment of Serum Response Factor and Hyperacetylation of Histones at Smooth Muscle-Specific Regulatory Regions During Differentiation of a Novel P19-Derived In Vitro Smooth Muscle Differentiation System Circ. Res., June 8, 2001; 88(11): 1127 - 1134. [Abstract] [Full Text] [PDF]
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M. D. Layne, S.-F. Yet, K. Maemura, C.-M. Hsieh, X. Liu, B. Ith, M.-E. Lee, and M. A. Perrella Characterization of the Mouse Aortic Carboxypeptidase-Like Protein Promoter Reveals Activity in Differentiated and Dedifferentiated Vascular Smooth Muscle Cells Circ. Res., April 5, 2002; 90(6): 728 - 736. [Abstract] [Full Text] [PDF]
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P. Qiu and L. Li Histone Acetylation and Recruitment of Serum Responsive Factor and CREB-Binding Protein Onto SM22 Promoter During SM22 Gene Expression Circ. Res., May 3, 2002; 90(8): 858 - 865. [Abstract] [Full Text] [PDF]
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