D-MEF2 is a MADS domain transcription factor expressed in the cardiac, somatic, and visceral muscle cell lineages in the Drosophila embryo. Genetic studies have demonstrated that D-mef2 gene function is required for the proper differentiation of all three of these muscle types. We show that D-MEF2 is also expressed in a limited number of other cells types during development, including Kenyon cells present in the mushroom bodies of the Drosophila brain. This finding suggests a role for D-mef2 in neuron differentiation. To investigate D-mef2 expression in muscle and Kenyon cells, we assayed 26 kb of D-mef2 5'-flanking and intragenic DNA for regulatory sequences controlling the expression of the gene. Our results show that separable enhancer sequences direct D-mef2 gene expression in the myogenic and neuronal cell lineages. The identification of these regulatory DNAs provides a starting point for the analysis of transcriptional regulators controlling the cell-specific expression of D-mef2 and a means to address the function of D-mef2 in Kenyon cell differentiation.