Thèse de doctorat
Résumé : During embryonic development, multipotent cardiovascular progenitor cells (MCPs) are specified from early mesoderm. Although the core cardiac transcriptional machinery acting during cardiac cell differentiation is relatively well known, the molecular mechanism acting upstream of these cardiac transcriptional factors, and promoting cardiac progenitor specification from early mesoderm remains poorly understood. We used embryonic stem cell (ESC) differentiation as a model to dissect the molecular mechanisms implicated in cardiovascular progenitor specification. Using ESCs, in which gene expression can be temporally regulated, we showed that transient expression of Mesp1 dramatically accelerates and enhances multipotent cardiovascular progenitor specification through an intrinsic and cellular autonomous mechanism. Using genome wide transcriptional analysis, we found that Mesp1 rapidly activates and represses a discrete set of genes. Using chromatin immunoprecipitation, we showed that Mesp1 directly binds to regulatory DNA sequences located in the promoter of many key genes belonging to the core cardiac transcriptional machinery, resulting in their rapid upregulation. Mesp1 also directly and strongly represses the expression of key genes regulating other early mesoderm and endoderm cell fates. Using engineered ESC expressing the green fluorescent protein under the control of the Mesp1 promoter, we isolated Mesp1 expressing cells in differentiating ESCs allowing characterization of the cellular and molecular mechanisms underlying cardiovascular specification. Our results demonstrate that Mesp1 acts as a key regulatory switch during cardiovascular specification, residing at the top of the hierarchy of the gene network responsible for cardiovascular cell fate determination. Moreover our results place Mesp1 upstream of the specification of both first and second heart fields and provide novel and important insights into the molecular mechanisms underlying the earliest step of cardiovascular specification. We identified cell surface markers expressed allowing the isolation of early cardiovascular progenitors and provide potentially novel methods for dramatically increasing the number of cardiovascular cells for cellular therapy in humans.