Function of the two xenopus Smad4s in early frog development

Signals from the transforming growth factor beta family members are transmitted in the cell through specific receptor-activated Smads and a common partner Smad4. Two Smad4 genes (alpha and beta/10, or smad4 and smad4.2) have been isolated from Xenopus, and conflicting data are reported for Smad4 beta/10 actions in mesodermal and neural induction. To further understand the functions of the Smad4s in early frog development, we analyzed their activities in detail. We report that Smad10 is a mutant form of Smad4 beta that harbors a missense mutation of a conserved arginine to histidine in the MH1 domain. The mutation results in enhanced association of Smad10 with the nuclear transcription corepressor Ski and leads to its neural inducing activity through inhibition of bone morphogenetic protein (BMP) signaling. In contrast to Smad10, both Smad4 alpha and Smad4 beta enhanced BMP signals in ectodermal explants. Using antisense morpholino oligonucleotides (MOs) to knockdown endogenous Smad4 protein levels, we discovered that Smad4 beta was required for both activin- and BMP-mediated mesodermal induction in animal caps, whereas Smad4 beta affected only the BMP signals. Neither Smad4 was involved directly in neural induction. Expression of Smad4 beta-MO in early frog embryos resulted in reduction of mesodermal markers and defects in axial structures, which were rescued by either Smad4 alpha or Smad4 beta. Smad4 alpha-MO induced only minor deficiency at late stages. As Smad4 beta, but not Smad4 alpha, is expressed at high levels maternally and during early gastrulation, our data suggest that although Smad4 alpha and Smad4 beta may have similar activities, they are differentially utilized during frog embryogenesis, with only Smad4 beta being essential for mesoderm induction.