Genetic interactions in zebrafish midline development

Mutational analyses have shown that the genes no tail (nt1, Brachyury homolog), floating head (flh, a Not homeobox gene), and cyclops (cyc) play direct and essential roles in the development of midline structures in the zebrafish. In both nt1 and flh mutants a notochord does not develop, and in cyc mutants the floor plate is nearly entirely missing. We made double mutants to learn how these genes might interact. Midline development is disrupted to a greater extent in cyc;flh double mutants than in either cyc or flh single mutants; their effects appear additive. Both the notochord and floor plate are completely lacking, and other phenotypic disturbances suggest that midline signaling functions are severely reduced. On the other hand, trunk midline defects in flh;nt1 double mutants are not additive, but are most often similar to those in nt1 single mutants. This finding reveals that loss of nt1 function can suppress phenotypic defects due to mutation at flh, and we interpret it to mean that the wild-type allele of nt1 (nt1⁺) functions upstream to flh in a regulatory hierarchy. Loss of function of nt1 also strongly suppresses the floor plate deficiency in cyc mutants, for we found trunk floor plate to be present in cyc;nt1 double mutants. From these findings we propose that nt1⁺ plays an early role in cell fate choice at the dorsal midline, mediated by the Nt1 protein acting to antagonize floor plate development as well as to promote notochord development.