EXPRESSION OF NEURONAL ACETYLCHOLINE-RECEPTOR GENES IN VERTEBRATE SKELETAL-MUSCLE DURING DEVELOPMENT

Of the 15 nicotinic ACh receptor genes identified in vertebrates, only four (alpha 1, beta 1, gamma, and delta) have been shown to be expres sed in embryonic skeletal muscle at early times. In mammalian muscle a fifth gene (epsilon) replaces the gamma gene in expression at later t imes. The remaining 10 nicotinic receptor genes identified to date (al pha 2-alpha 8, beta 2-beta 4) are expressed in the nervous system and are considered neuronal genes. Using RNase protection assays, we shaw here that four of the neuronal-type genes (alpha 4, alpha 5, alpha 7, and beta 4) are expressed in developing chick skeletal muscle. Two of them (alpha 4 and alpha 7) decline substantially in transcript abundan ce between embryonic days 11 and 17, as does alpha 1, while the other two (alpha 5 and beta 4) show only moderate decreases over the same ti me period. At embryonic day 8, alpha 7 transcripts are nearly 20% as a bundant as alpha 1 transcripts. In site hybridizations confirm the pre sence of alpha 7 transcripts in muscle cells both in cell culture and in embryonic tissue. No evidence was found for expression of the alpha 2, alpha 3, alpha 8, or beta 3 genes in muscle. Immunoprecipitations and immunoblot analysis using subunit-specific monoclonal antibodies r eveal alpha 7 protein in muscle, and the amount of protein rises and d eclines with the amount of alpha 7 mRNA during development. Sucrose gr adient analysis demonstrates that the alpha 7 protein is present in mu scle as a species of 10S, the size expected for a nicotinic receptor. The alpha 7 species in muscle binds alpha-bungarotoxin but does not co ntain alpha 1 subunits, indicating that the two kinds of alpha-type ge ne products segregate during assembly. The results suggest that neuron al AChRs may play a role in early muscle development.