Expression and regulation of eHAND during limb development.

eHAND is a bHLH transcription factor with important functions during embryogenesis. Here, we report that eHAND has a dynamic pattern of expression during limb development. In chick embryos, eHAND expression is first observed in the ventral mesoderm of the emerging limb. Its expression is then restricted to an anteroventral area of mesoderm at mid-level in the proximodistal axis. At later stages, expression is observed in the autopod encompassing the ventral tendons of the digits. In mouse embryos, only the anteroventral domain of expression is conserved, the early ventral expression not being detectable and the late pattern of expression differing clearly from that in the chick. A constant feature of all areas of expression is their ventral and anterior localization. Respecification of the anterior mesoderm as occurs secondarily to Sonic hedgehog (SHH) or retinoic acid application to the anterior border leads to down-regulation of eHAND expression. Accordingly, eHAND expression is not detectable in talpid(2) mutant limbs, which are considered to be posteriorized limbs. However, eHAND expression is little modified in oligozeugodactyly, a chick mutant that lacks Shh signaling in the limb but retains certain anteroposterior polarity. Interestingly, eHAND expression is also linked to the ventral identity of the mesoderm and is repressed by the dorsal ectoderm. It is also positively regulated by bone morphogenetic protein signaling, which is also known to participate in dorsoventral patterning. We suggest that eHAND expression may be related to the anteroventral identity of the mesoderm. However, in overexpression experiments using retroviral vectors, only a low percentage of cases (5%) showed phenotypic alterations, consisting of a duplication of digit 2.