The Limb Bud Shh-Fgf Feedback Loop Is Terminated by Expansion of Former ZPA Cells

Vertebrate limb outgrowth is driven by a positive feedback loop involving Sonic Hedgehog ( Shh ), Gremlin , and Fgf4 . By overexpressing individual components of the loop at a time after these genes are normally down-regulated in chicken embryos, we found that Shh no longer maintains Gremlin in the posterior limb. Shh -expressing cells and their descendants cannot express Gremlin . The proliferation of these descendants forms a barrier separating the Shh signal from Gremlin -expressing cells, which breaks down the Shh-Fgf4 loop and thereby affects limb size and provides a mechanism explaining regulative properties of the limb bud.