Sonic Hedgehog (SHH) is a fundamental zone of polarizing activity (ZPA) morphogen well characterized for a key role in vertebrate limb patterning. In amphibians, however, its function remains incompletely understood regarding tetrapod evolution. Leveraging the allotetraploid genome of Xenopus laevis, we demonstrate a clear functional and regulatory divergence between the two shh homeologs (L and S). Live imaging of a locus-targeted egfp reporters revealed that only shh.L is active in the limb ZPA, while both genes are active in trunk tissues. Crucially, CRISPR-Cas9-mediated disruption of shh.L shows its role extends beyond distal patterning: its ablation results in the complete absence of limb initiation. This limbless phenotype, contrasting with truncated limbs in amniotes, identifies shh as a key trigger for anuran limb formation and indicates evolutionary diversification of developmental programs across tetrapods.
Kagawa, N., Mizuno, R., Umesono, Y., Suzuki, K.-i. T., Mochii, M.
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