The mouse homeodomain protein Phox2 regulates Ncam promoter activity in concert with Cux/CDP and is a putative determinant of neurotransmitter phenotype.

Transcriptional regulation of the gene encoding the cell adhesion receptor NCAM (neural cell adhesion molecule), a putative effector molecule of a variety of morphogenetic events, is likely to involve important regulators of morphogenesis. Here we identify two mouse homeodomain proteins that bind to an upstream regulatory element in the Ncam promoter: Cux, related to Drosophila cut and human CDP, and Phox2, a novel protein with a homeodomain related to that of the Drosophila paired gene. In transient transfection experiments, Cux was found to be a strong inhibitor of Ncam promoter activity, and this inhibition could be relieved by simultaneously overexpressing Phox2. These results suggest that the Ncam gene might be a direct target of homeodomain proteins and provide a striking example of regulatory cross-talk between homeodomain proteins of different classes. Whereas the expression pattern of Cux/CDP includes many NCAM-negative sites, Phox2 expression was restricted to cells also expressing Ncam or their progenitors. The localisation data thus strongly reinforce the notion that Phox2 plays a role in transcriptional activation of Ncam in Phox2-positive cell types. In the peripheral nervous system, Phox2 was strongly expressed in all ganglia of the autonomic nervous system and more weakly in some cranial sensory ganglia, but not in the sensory ganglia of the trunk. Phox2 transcripts were detected in the primordia of sympathetic ganglia as soon as they form. Phox2 expression in the brain was confined to spatially restricted domains in the hindbrain, which correspond to the noradrenergic and adrenergic nuclei once they are identifiable. All Phox2-expressing components of the peripheral nervous system are at least transiently adrenergic or noradrenergic. In the developing brain, Phox2 was expressed at all known locations of (nor)adrenergic neurones and of their precursors. These results suggest that Phox2, in addition to regulating the NCAM gene, may be part of the regulatory cascade that controls the differentiation of neurons towards this neurotransmitter phenotype.