Nucleic Acids Research, 1992, Vol. 20, No. 7 1793-1799
© 1992
MOLECULAR BIOLOGY |
Cis-acting elements responsible for muscle-specific expression of the myosin heavy chain ß gene
1Departments of Medicine, The University of Chicago Chicago, IL 60637 2Departments of Organismal Biology and Anatomy, The University of Chicago Chicago, IL 60637 3Departments of Pharmacological & Physiological Sciences, The University of Chicago Chicago, IL 60637 4Marine Biology Laboratory Woods Hole, MA 02543 5Department of Medicine, University of Alabama at Birmingham Birmingham, AL 35294, USA
*To whom correspondence should be addressed at Department of Medicine, The University of Chicago, Hospital Box 360, 5841 S. Maryland Avenue, Chicago, IL 60637, USA
Received August 8, 1991. Revised February 19, 1992. Accepted February 19, 1992.
The 5' flanking region of the rabbit myosin heavy chain (HC) ß gene extending 295 bp upstream from the cap site provides muscle-specific transciiptional activity. In this study, we have identified and functionally characterized cls-acting elements that regulate the muscle-specific expression within this region. By using linker-scanner (LS) mutants between - 295 bp and a putative TATA box, we found five distinct positive cis-acting sequences necessary for transcription: element A, the sequences between –276 and –263, which contains a putative M-CAT motif in an inverted orientation; B, the sequences between –207 and –180; C, the sequences between –136 and –127; D, the sequences between –91 and –80; and E, a TATA consensus sequence at –28. The fragment containing both A and B elements dramatically enhanced the expression of the chloramphenicol acetyltransferase (CAT) gene driven by a heteroiogous promoter in differentiated muscle cells, whereas fragments containing either A or B elements alone had little or no effect in either muscle or nonmuscle cells. Therefore, these two elements appear to act cooperatively in determining a high level of muscle- and stage-specific expression. Unlike the typical enhancer element, this region functions in an orientation-dependent manner. In contrast, the fragment containing C and D elements activates the heteroiogous promoter in both muscle and nonmuscle cells in an orientation-independent manner.
Present address: Molecular Medicine Unit, Beth Israel Hospital, 330 Brooklin Avenue, Boston, MA 02215, USA
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