RAPsyn (also known as 43K protein), a mouse muscle protein localized to the synaptic membrane, is thought to be involved in the localization of nicotinic acetylcholine receptors at the neuromuscular junction. We have characterized the transcriptional regulation of the RAPsyn gene and the synthesis of the RAPsyn protein during muscle cell differentiation. Nuclear run-on experiments and RNAase protection analyses showed that mRNA encoding RAPsyn, but not the acetylcholine receptor subunits, is present in undifferentiated muscle cells. The RAPsyn protein present in undifferentiated and differentiated muscle cells cannot be distinguished by peptide maps, turnover rates, cellular subfractionation, or ability to incorporate myristate. Whereas the amount of acetylcholine receptor subunit mRNA is increased approximately 100-fold after denervation, the amount of RAPsyn mRNA is increased just 2- to 3-fold. We conclude that the expression of RAPsyn and the acetylcholine receptor is not coordinately regulated in mouse muscle.