The basic helix-loop-helix transcription factor Hand2, together with Ascl1, Phox2a, Phox2b and Gata2/Gata3, is induced by bone morphogenetic proteins in neural crest-derived precursor cells during sympathetic neuron generation. Hand2 overexpression experiments and the analysis of its function at the Dbh promotor implicated Hand2 in the control of noradrenergic gene expression. Using the zebrafish hand2 deletion mutant hands off, we have now investigated the physiological role of hand2 in the development of sympathetic ganglia. In hands off mutant embryos, sympathetic precursor cells aggregate to form normal sympathetic ganglion primordia characterized by the expression of phox2b, phox2a and the achaete-scute family member zash1a/ascl1. The expression of the noradrenergic marker genes th and dbh is strongly reduced, as well as the transcription factors gata2 and tfap2a (Ap-2 α). By contrast, generic neuronal differentiation seems to be unaffected, as the expression of elavl3 (HuC) is not reduced in hands off sympathetic ganglia. These results demonstrate in vivo an essential and selective function of hand2 for the noradrenergic differentiation of sympathetic neurons, and implicate tfap2a and gata2 as downstream effectors.
Neural crest cells give rise to numerous derivatives including chromatophores, craniofacial cartilages, and neurons and glia of the peripheral nervous system. Likewise, lateral plate mesoderm generates numerous blood, cardiac and vascular derivatives. hatchback (hbk) is an ENU-induced recessive embryonic lethal mutation. All chromatophores are absent or nearly absent in hbk mutants. Sympathetic, dorsal root ganglion and enteric neurons are also missing. The absence of neural crest-derivatives is preceded by abnormal expression of multiple transcription factors known to regulate early neural crest development. hbk mutant embryos also display defects in cardiovascular development as evidenced by cardiac edema and a lack of blood and circulation. Abnormal expression of several genes known to regulate the development of cardiac, vascular and hematopoietic progenitors has also been observed. Neural plate and axial and paraxial mesoderm development in hbk mutants is indistinguishable from wild-type siblings. These observations suggest hbk function is specifically required for early neural crest development, as well as for the establishment and development of specific lateral plate mesoderm-derived lineages.