Hand is a direct target of the forkhead transcription factor Biniou during Drosophila visceral mesoderm differentiation

The visceral trunk mesoderm in Drosophila melanogaster develops under inductive signals from the ectoderm. This leads to the activation of the key regulators Tinman, Bagpipe and Biniou that are crucial for specification of the circular visceral muscles. How further differentiation is regulated is widely unknown, therefore it seems to be essential to identify downstream target genes of the early key regulators. In our report we focus on the analysis of the transcriptional control of the highly conserved transcription factor Hand in circular visceral muscle cells, providing evidence that the hand gene is a direct target of Biniou.

In the present report we show that cross species sequence comparison of non-coding sequences between orthologous genes is a powerful tool to identify conserved regulatory elements. Combining functional dissection experiments in vivo and protein/DNA binding studies we identified hand as a direct target of Biniou in the circular visceral muscles.

Herein we describe the identification of a regulatory region in the hand gene essential and sufficient for the expression in the visceral mesoderm during embryogenesis. We found that hand expression in the circular visceral mesoderm is abolished in embryos mutant for the FoxF domain containing transcription factor Biniou. Furthermore we demonstrate that Biniou regulates hand expression by direct binding to a 300 bp sequence element, located within the 3rd intron of the hand gene. This regulatory element is highly conserved in different Drosophila species. In addition, we provide evidence that Hand is dispensable for the initial differentiation of the embryonic visceral mesoderm.