Hoxc genes play a central role in claw/nail development

Abstract

In amniotes, Hox genes are found in 4 clusters, two of them, the HoxA and HoxD clusters, play remarkable roles in limb development. In contrast, the HoxB or HoxC clusters seem to be practically dispensable for proper limb growth and skeletal patterning. Recently, we turned our attention to Hoxc genes as the temporal transcriptome of the isolated limb components (mesoderm and ectoderm) exposed a collinear activation of the HoxC cluster specifically in the limb ectoderm. In situ hybridization supported this temporal colinear activation and showed a progressive restriction of the expression of more 5' HoxC members to the dorsal side of thedigit tip coincident with the claw/nail region. The histological an immunohistochemical reevaluation of mice lacking the complete HoxC cluster showed the absence of the nail organ. To identify the cis-regulatory mechanism controlling Hoxc gene transcription in the limb ectoderm, we used the ATACseq technique. These experiments revealed two open chromatin region located 5' to the HoxC cluster, termed EC1 and EC2, that were highly conserved in mammals and in placentaria, respectively. These two regulatory regions were able to elicit reporter gene activity in the limb ectoderm in mouse transgenic assays, although the CRISPR-mediated individual deletion of either EC1 or EC2 had no phenotypic consequence. To further clarify the role of EC1 and EC2, we are currently generating the CRISPR-mediated combined deletion of both putative enhancer regions. Our study sheds light on the HoxC dependent control on nail/claw/hoof formation at the tip of the digits. The form and size of this epidermal derivative always correlates with that of the distal phalanx showing together a wide range of adaptive strategies across tetrapod species.