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J. Biol. Chem., Vol. 278, Issue 46, 45435-45444, November 14, 2003

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Muscle Induces Neuronal Expression of Acetylcholinesterase in Neuron-Muscle Co-culture

TRANSCRIPTIONAL REGULATION MEDIATED BY cAMP-DEPENDENT SIGNALING^*

Joy X. S. Jiang, Roy C. Y. Choi, Nina L. Siow, Henry H. C. Lee, David C. C. Wan¶, and Karl W. K. Tsim||

From the Department of Biology and Molecular Neuroscience Center, The Hong Kong University of Science and Technology, Clear Water Bay Road, Kowloon, Hong Kong, China and the ^¶Department of Biochemistry, The Chinese University of Hong Kong, Shatin, Hong Kong, China

Presynaptic motor neuron synthesizes and secretes acetylcholinesterase (AChE) at vertebrate neuromuscular junctions. In order to determine the retrograde role of muscle in regulating the expression of AChE in motor neuron, a chimeric co-culture of NG108-15 cell, a cholinergic cell line that resembles motor neuron, with chick myotube was established to mimic the neuromuscular contact in vitro. A DNA construct of human AChE promoter tagged with luciferase (pAChE-Luc) was stably transfected into NG108-15 cells. The co-culture with myotubes robustly stimulated the promoter activity as well as the endogenous expression of AChE in pAChE-Luc stably transfected NG108-15 cells. Muscle extract derived from chick embryos when applied onto pAChE-Luc-expressing NG108-15 cells induced expressions of AChE promoter and endogenous AChE. The cAMP-responsive element mutation on human AChE promoter blocked the muscle-induced AChE transcriptional activity in cultured NG108-15 cells either in co-culturing with myotube or in applying muscle extract. The accumulation of intracellular cAMP and the phosphorylation of cAMP-responsive element-binding protein in cultured NG108-15 cells were stimulated by applied muscle extract. Part of the muscle-induced signaling was mimicked by application of calcitonin gene-related peptide in cultured NG108-15 cells. These results suggest the muscle-induced neuronal AChE expression in the co-culture is mediated by a cAMP-dependent signaling.

Received for publication, June 16, 2003 , and in revised form, September 3, 2003.

^* This work was supported by Research Grants Council of Hong Kong Grants 6098/02M (to K. W. K. T.) and 4140/98M (to D. C. C. W.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Supported by the postdoctoral matching fund from Hong Kong University of Science and Technology.

^|| To whom correspondence should be addressed. Tel.: 852-2358-7332; Fax: 852-2358-1559; E-mail: BOTSIM{at}UST.HK.

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