We have identified a mesoderm-specific Drosophila gene, designated D-mef2. The encoded protein contains the MADS- and MEF2-specific domains, which are characteristic of the myocyte-specific enhancer factor 2 (MEF2) family of transcription factors. D-mef2 RNA is first detectable in the presumptive mesoderm at late cellular blastoderm stage and is expressed in all mesoderm after invagination. Following the dorsal migration of the mesodermal layer, D-mef2 expression becomes restricted to the primordia for visceral muscle and the heart. In the second phase, D-mef2 expression is first distinct in heart precursors and then becomes prominent sequentially in visceral and somatic muscles. twi activity is required for D-mef2 expression, while sna function may be needed for the maintenance of D-mef2 expression but not its initiation. D-mef expression is not dependent on the function of tin, and embryos that are deficient for the mesodermal gene DFR1 also show normal initiation of D-mef2 expression at blastoderm. These results suggest that D-mef2 could have a function in early mesoderm differentiation and may be required for subsequent cell fate specifications within the somatic and visceral/heart mesodermal layers.