MicroRNA-92a/Sp1/MyoD axis regulates hypoxic stimulation of myogenic-lineage differentiation in mouse embryonic stem cells

Abstract Hypoxic microenvironments exist in developing embryonic tissues and determine stem cell fate. We previously demonstrated that hypoxic priming plays roles in lineage commitment of embryonic stem cells. In the present study, we found that hypoxia-primed embryoid bodies (Hyp-EBs) efficiently differentiate into the myogenic lineage, resulting in the induction of the myogenic marker, MyoD, which was not mediated by HIF1α or HIF2α, but rather by Sp1 induction and binding to the MyoD promoter. Knock-down of Sp1 in Hyp-EBs abrogated hypoxia-induced MyoD expression and myogenic differentiation. Importantly, in the cardiotoxin-muscle injury mice model, Hyp-EB transplantation facilitated muscle regeneration in vivo, whereas transplantation of Sp1-knock-down Hyp-EBs failed to do. Moreover, we compared miRNA expression profiles between EBs under normoxia versus hypoxia, and found that hypoxia-mediated Sp1 induction was mediated by the suppression of microRNA-92a, which directly targeted the 3′-untranslated region (3′-UTR) of Sp1. Further, the inhibitory effect of microRNA-92a on Sp1 in luciferase assay was abolished by a point mutation in specific sequence in the Sp1 3′-UTR that is required for the binding of microRNA-92a. Collectively, these results suggest that hypoxic priming enhances EB commitment to the myogenic lineage through miR-92a/Sp1/MyoD regulatory axis, suggesting a new pathway that promotes myogenic-lineage differentiation.