BMP ligands interact with two types of BMP receptors (BMPR1 and BMPR2), which in turn trigger intracellular signal transduction by phosphorylation of the receptor-associated SMAD1, -5 and -8 proteins. In summary, our analysis reveals that SMAD4 is required to initiate: (1) formation of the Sox9-positive digit ray primordia and (2) aggregation and chondrogenic differentiation of all limb skeletal elements.īone morphogenetic proteins (BMPs) are ligands belonging to the transforming growth factor beta (TGFβ) superfamily, which control inductive processes in early embryos and during organogenesis (reviewed by Zakin and De Robertis, 2010) and are required for chondrogenesis, growth and ossification of bones in vertebrates (reviewed by Wu et al., 2007). Conditional inactivation of Bmp2 and Bmp4 indicated that the loss of digit ray primordia and increase in connective tissue were predominantly a consequence of disrupting SMAD4-mediated BMP signal transduction. This pointed to a general loss of tissue organization and diversion of mutant cells toward non-specific connective tissue. The progressive loss of SOX9 due to disrupting digit ray primordia and chondrogenesis was paralleled by alterations in genes marking other lineages. At the cellular level, Smad4 deficiency blocked the aggregation of Sox9-positive progenitors, thereby preventing chondrogenic differentiation as revealed by absence of collagen type II. Specific inactivation of Smad4 during handplate development pointed to its differential requirement for posterior and anterior digit ray primordia. While this Smad4 inactivation did not alter the early Sox9 distribution, prefiguring the chondrogenic primordia of the stylopod and zeugopod, it disrupted formation of all Sox9-positive digit ray primordia. SMAD4 is an essential mediator of canonical TGFβ/BMP signal transduction and we inactivated Smad4 in mouse limb buds from early stages onward to study its functions in the mesenchyme.

Primordia resolution