Functional analysis of a first hindlimb positioning enhancer via expression.

During the early development of tetrapods, including humans, the embryonic body elongates caudally once the anterior-posterior axis is established. During this process, region-specific vertebral morphogenesis occurs, with the determination of limb positioning along the anterior-posterior axis. We previously reported that functions as an anatomical integration system that determines the positioning of hindlimbs and sacral vertebrae where is expressed. However, the molecular mechanisms underlying induction of expression remain unclear. In this study, we searched for non-coding regions near the locus that were conserved across species to elucidate the regulatory mechanisms of expression. We identified an enhancer of the gene in intron 1 and named it highly conserved region (HCR). In HCR knockout mice, the expression level of endogenous was decreased, and the position of the sacral-hindlimb unit was shifted posteriorly. We also searched for factors upstream of based on the predicted transcription factor binding sites within the HCR. We found that inhibition of FGF signaling increased endogenous expression, suggesting that FGF signaling negatively regulates expression. However, FGF signaling does not regulate HCR activity. Our results suggest that there are species-specific enhancers other than HCR and that FGF signaling regulates expression independent of HCR.