TGF-β signaling and Creb5 cooperatively regulate Fgf18 to control pharyngeal muscle development

The communication between myogenic cells and their surrounding connective tissues is indispensable for muscle morphogenesis. During late embryonic development in mice, myogenic progenitors migrate to discrete sites to form individual muscles. The detailed mechanism of this process remains unclear. Using levator veli palatini (LVP) development as a model, we systematically investigated how a distinct connective tissue subpopulation, perimysial fibroblasts, communcates with myogenic cells to regulate mouse pharyngeal myogenesis. Using single-cell RNAseq data analysis, we identified that TGF-β signaling is a key regulator for the perimysial fibroblasts. Loss of TGF-β signaling led to defects in perimysial fibroblasts and subsequently muscle formation in Osr2-Cre;Alk5^fl/fl mice. In particular, a perimysial fibroblast-specific regulator, Creb5, interacts with TGF-β signaling to enable specific activation of perimysial fibroblast-derived signals such as Fgf18. Moreover, Fgf18 supports pharyngeal muscle development in vivo and its exogenous expression can partially rescue myogenic cell numbers in Osr2-Cre;Alk5^fl/fl samples, illustrating that TGF-β-regulated Fgf18 signaling is required for LVP development. Collectively, our findings revealed the mechanism by which TGF-β signaling achieves its specificity in defining the perimysial-to-myogenic signals for pharyngeal myogenesis.