Abstract
ABSTRACTThe key molecular interactions governing vertebrate limb bud development are a paradigm for studying the mechanisms controlling progenitor cell proliferation and specification during vertebrate organogenesis. However, little is known about the cellular heterogeneity of the mesenchymal progenitors in early limb buds that ultimately contribute to the chondrogenic condensations prefiguring the skeleton. We combined flow cytometric and transcriptome analyses to identify the molecular signatures of several distinct mesenchymal progenitor cell populations present in early mouse forelimb buds. In particular, jagged 1 (JAG1)-positive cells located in the posterior-distal mesenchyme were identified as the most immature limb bud mesenchymal progenitors (LMPs), which crucially depend on SHH and FGF signaling in culture. The analysis of gremlin 1 (Grem1)-deficient forelimb buds showed that JAG1-expressing LMPs are protected from apoptosis by GREM1-mediated BMP antagonism. At the same stage, the osteo-chondrogenic progenitors (OCPs) located in the core mesenchyme are already actively responding to BMP signaling. This analysis sheds light on the cellular heterogeneity of the early mouse limb bud mesenchyme and on the distinct response of LMPs and OCPs to morphogen signaling.
Highlights
The developing vertebrate limb bud is an excellent model for studying the molecular mechanisms and cellular interactions that govern proliferative expansion, specification and differentiation of mesenchymal progenitors during organogenesis
A feedback signaling system is established between the posterior SHH signaling center and FGF signaling by the apical ectodermal ridge (AER), which regulates the survival and proliferative expansion of limb bud mesenchymal progenitors (LMPs) in concert with gremlin 1 (GREM1)-mediated BMP antagonism and WNT signaling
Among the live Lin− mesenchymal cells, the fraction of S9−jagged 1 (JAG1)+ LMPs was reduced by ∼42%, while S9−Pαhi LMPs were not significantly affected and the fraction of the predominant S9+Pαhi osteo-chondrogenic progenitors (OCPs) increased by ∼15% in gremlin 1 (Grem1)-deficient forelimb buds (Fig. 6E). These results show that GREM1-mediated BMP antagonism (Zuniga et al, 1999) preferentially impacts the immature S9−JAG1+ LMPs located in the distal-posterior forelimb bud mesenchyme. This analysis highlighted the importance of GREM1-mediated protection of LMPs from premature exposure to BMP signaling in early forelimb buds
Summary
The developing vertebrate limb bud is an excellent model for studying the molecular mechanisms and cellular interactions that govern proliferative expansion, specification and differentiation of mesenchymal progenitors during organogenesis. Muscles arise from myoblasts that migrate from the somites into the early limb bud It has been shown that SHH morphogen signaling specifies the anteroposterior (AP) identities of the future digits during the onset of mouse limb bud outgrowth (around embryonic day E9.75-E10.5; Zhu et al, 2008). Much less is known about the cellular heterogeneity of the mesenchyme and potential differences in the mesenchymal response to morphogen signaling
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