We describe the dissection of a body muscle-specific enhancer sequence contained within the Caenorhabditis elegans myosin heavy chain gene unc-54. A 90-base pair segment that was sufficient for both enhancer function and tissue specificity was subjected to mutational analysis. Several separated sites within this region were required for activity; mutations in these sites led to dramatic decreases in enhancer activity, while substitutions in the intervening regions had minimal effects on activity. The individual sites appear to function as semi-independent and partially interchangeable enhancer subelements, as seen by our ability to create functional enhancers by constructing novel multimers and combinations. Four different enhancer subelements (designated O, I, II, and III) were identified in this way. Although partially interchangeable, some differences between these subelements were evident. In particular, concatamers of site III exhibited the highest levels of activity but had a broader tissue specificity than the intact enhancer, including both hypodermal and muscle tissue. The specificity of the intact enhancer thus reflects a combinatorial function of the specificities of the constituent subelements.