Regulation of the tinman homologues in Xenopus embryos

Vertebrate homologues of the Drosophila tinman transcription factor have be en implicated in the processes of specification and differentiation of card iac mesoderm. In Xenopus three members of this family have been isolated to date. Here we show that the XNkx2-3, XNkx2-5, and XNkx2-10 genes are expre ssed in increasingly distinctive patterns in endodermal and mesodermal germ layers through early development, suggesting that their protein products ( either individually or in different combinations) perform distinct function s. Using amphibian transgenesis, we find that the expression pattern of one of these genes, XNkx2-5, can be reproduced using transgenes containing onl y 4.3 kb of promoter sequence. Sequence analysis reveals remarkable conserv ation between the distalmost 300 bp of the Xenopus promoter and a portion o f the AR2 element upstream of the mouse and human Nkx2-5 genes. Interesting ly, only the 3' half of this evolutionarily conserved sequence element is r equired for correct transgene expression in frog embryos. Mutation of conse rved GATA sites or a motif resembling the dpp-response element in the Droso phila tinman tinD enhancer dramatically reduces the levels of transgene exp ression. Finally we show that, despite its activity in Xenopus embryos, in transgenic mice the Xenopus Nkx2-5 promoter is able to drive reporter gene expression only in a limited subset of cells expressing the endogenous gene . This intriguing result suggests that despite evolutionary conservation of some cia-regulatory sequences, the regulatory controls on Nkx2-5 expressio n have diverged between mammals and amphibians. (C) 2000 Academic Press.