Interlinked signaling feedback loops and self-regulation during vertebrate limb development

Reliable organ and tissue morphogenesis seems to depend on self-regulatory mechanisms that are able to buffer spontaneous and/or genetic variations. While the analysis of the interactions of BMPs with their antagonists during gastrulation has provided insights into the self-regulatory capacity of early embryos, few other mechanisms endowing developmental programmes with robustness have been identified. Limb development and digit specification are regulated by epithelial-mesenchymal (e-m) interactions involving instructive SHH and FGF signaling. The BMP antagonist Gremlin1 (GREM1) is key to establishing these interactions. By combining genetics with ex vivo manipulation and mathematical modeling, we establish that both BMP4 and SHH positively regulate Grem1 expression, defining this antagonist as a critical node that links a fast and self-regulatory BMP4/GREM1 module to the SHH/GREM1/FGF e-m feedback loop. This network architecture allows a self-regulative switch from BMP4- to SHH-driven limb development and endows limb patterning with robustness against variations in gene activity due to interconnectivity between the BMP, SHH and FGF signaling pathways. The establishment of this robust signaling network may have contributed to the appearance and stabilization of pentadactyly in tetrapods.