Multi-modal effects of BMP signaling on Nodal expression in the lateral plate mesoderm during left-right axis formation in the chick embryo.

During development of left-right asymmetry in the vertebrate embryo, Nodal plays a central role for determination of left-handedness. Bone morphogenetic protein (BMP) signaling has an important role for regulation of Nodal expression, although there is controversy over whether BMP signaling has a positive or negative effect on Nodal expression in the chick embryo. As BMP is a morphogen, we speculated that different concentrations might induce different responses in the cells of the lateral plate mesoderm (LPM). To test this hypothesis, we analyzed the effects of various concentrations of BMP4 and NOGGIN on Nodal expression in the LPM. We found that the effect on Nodal expression varied in a complex fashion with the concentration of BMP. In agreement with previous reports, we found that a high level of BMP signaling induced Nodal expression in the LPM, whereas a low level inhibited expression. However, a high intermediate level of BMP signaling was found to suppress Nodal expression in the left LPM, whereas a low intermediate level induced Nodal expression in the right LPM. Thus, the high and the low intermediate levels of BMP signaling up-regulated Nodal expression, but the high intermediate and low levels of BMP signaling down-regulated Nodal expression. Next, we sought to identify the mechanisms of this complex regulation of Nodal expression by BMP signaling. At the low intermediate level of BMP signaling, regulation depended on a NODAL positive-feedback loop suggesting the possibility of crosstalk between BMP and NODAL signaling. Overexpression of a constitutively active BMP receptor, a constitutively active ACTIVIN/NODAL receptor and SMAD4 indicated that SMAD1 and SMAD2 competed for binding to SMAD4 in the cells of the LPM. Nodal regulation by the high and low levels of BMP signaling was dependent on Cfc up-regulation or down-regulation, respectively. We propose a model for the variable effects of BMP signaling on Nodal expression in which different levels of BMP signaling regulate Nodal expression by a balance between BMP-pSMAD1/4 signaling and NODAL-pSMAD2/4 signaling.