Functional interactions between Drosophila bHLH/PAS, Sox, and POU transcription factors regulate CNS midline expression of the slit gene

During Drosophila embryogenesis the CNS midline cells have organizing activ ities that are required for proper elaboration of the axon scaffold and dif ferentiation of neighboring neuroectodermal and mesodermal cells. CNS midli ne development is dependent on Single-minded (Sim), a basic-helix-loop-heli x (bHLH)-PAS transcription factor. We show here that Fish-hook (Fish), a So x HMG domain protein, and Drifter (Dfr), a POU domain protein, act in conce rt with Single-minded to control midline gene expression. single-minded, fi sh-hook, and drifter are all expressed in developing midline cells, and bot h loss- and gain-of-function assays revealed genetic interactions between t hese genes. The corresponding proteins bind to DNA sites present in a 1 kb midline enhancer from the slit gene and regulate the activity of this enhan cer in cultured Drosophila Schneider line 2 cells. Fish-hook directly assoc iates with the PAS domain of Single-minded and the POU domain of Drifter; t he three proteins can together form a ternary complex in yeast. In addition , Fish can form homodimers and also associates with other bHLH-PAS and POU proteins. These results indicate that midline gene regulation involves the coordinate functions of three distinct types of transcription factors. Func tional interactions between members of these protein families may be import ant for numerous developmental and physiological processes.