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Development, Vol 115, Issue 1 213-219, Copyright © 1992 by Company of Biologists
JOURNAL ARTICLES |
KG Chahine, W Walke and D Goldman
Mental Health Research Institute, University of Michigan, Ann Arbor 48109.
Muscle electrical activity suppresses expression of the embryonic-type (alpha, beta, gamma, and delta) nicotinic acetylcholine receptor (nAChR) genes. The molecular mechanism by which electrical activity regulates these genes is not known. One approach to this problem is to identify regions of the nAChR genes that mediate electrical regulation. Here we report results from such a study of the nAChR delta-subunit gene. We cloned the rat delta-subunit promoter region and created an expression vector in which this DNA controlled the expression of a down-stream luciferase structural gene. The effect that muscle electrical activity had on the expression from this promoter was assayed by introducing this expression vector into electrically stimulated and tetrodotoxin (TTX)-treated rat primary myotubes, and assaying for luciferase activity. These myotubes, when stimulated with extracellular electrodes, suppressed endogenous embryonic-type nAChR gene expression compared to those treated with TTX. Transfection of these cells with delta-promoter-luciferase expression vectors resulted in the delta-promoter conferring electrical regulation on luciferase expression. Additional experiments using deletions from the 5' end of the delta-promoter region have identified an element between -677 and -550 bp that suppressed delta-promoter activity and a minimal 102 bp sequence that promotes and regulates luciferase expression in response to muscle electrical activity. This latter sequence also contains all the necessary elements to confer tissue and developmental stage-specific expression.
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