DNA Sequences Mediating the Transcriptional Response of theMix.2Homeobox Gene to Mesoderm Induction

AbstractPeptide growth factors can initiate changes in cell fate inXenopusectodermal explants and induce the formation of mesoderm. Marker genes expressed in mesoderm allow the analysis of whether, or how much, induction has occurred, but do not tell us what molecules are involved in carrying out the response. In this report we describe the isolation of genomic and cDNA clones ofMix.2, a gene closely related to theXenopushomeobox geneMix.1, and demonstrate that the promoter of theMix.2 gene is responsive to mesoderm induction signals when linked to a CAT reporter and microinjected into developingXenopusembryos. Like the chromosomalMix.1 gene, microinjectedMix.2 gene plasmids respond to activin in the presence of cycloheximide in animal cap assays and also respond to the embryonic inductive signal in Nieuwkoop recombinants. The injected promoter does not respond to TGF-β2 or FGF. Deletion analysis of theMix.2 promoter demonstrated that sequences required for maximal transcriptional activity in response to mesoderm induction are scattered across a 290-bp region. This is the first report of a microinjected plasmid responding to immediate-early transcriptional activation in developingXenopusembryos. This assay reduces the complexity of the cellular response to embryonic induction to the simple question of which molecules activate theMix.2 promoter and provides a sensitive and rapid test with which to pursue the answer