The role of Tbx6 in regulating the differentiation of multipotent stem cells
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Stem cells hold great promise for the regeneration of injured tissues in a wide variety of important medical conditions. However, the molecular mechanisms that control differentiation remain poorly understood. The T-box (Tbx) family of transcription factors is integral to normal embryonic development. Abnormalities in Tbx expression/function result in defects such as DiGeorge/Velocardiofacial (Tbx1), ulnar-mammary (Tbx3), and Holt-Oram (Tbx5) syndromes. Tbx6 is essential for the specification of the posterior paraxial mesoderm and as knockout studies in mice have suggested, its absence directs posterior somites to differentiate along the default neural pathway. My hypothesis was that Tbx6 mediates the differentiation of multipotent cells to the cardiac myocyte lineage while suppressing differentiation into neurons/glia. To this end, I have used multipotent P19CL6 cell line which can undergo stimulus-specific cardiac myocyte or neuronal/glial differentiation.;I found that the abundance of Tbx6 protein was increased during DMSO-induced differentiation of P19CL6 cells to cardiac myocytes. Tbx6 overexpression was not sufficient by itself to induce the differentiation of P19CL6 cells to cardiac myocytes as assessed by expression of myocyte markers and spontaneous beating of the cultured cells. In contrast, knockdown of endogenous Tbx6 inhibited DMSO-induced differentiation of P19CL6 cells to cardiac myocytes as assessed by the above parameters, while ablating the expression of Nkx2.5, a homeobox protein that plays an early and critical role in cardiogenesis. In contrast to cardiac myocyte differentiation, the abundance of Tbx6 declined during the RA-induced differentiation of P19CL6 cells to neurons and glial cells. As suggested by the Tbx6 deficient murine model, Tbx6 knockdown potentiated RA-induced differentiation to neurons and glial cells. Conversely, Tbx6 overexpression interfered with differentiation in this paradigm.;In conclusion, Tbx6 is necessary but not sufficient for cardiac differentiation of P19CL6 cells, and may function as a molecular switch that promotes the differentiation of multipotent cells into cardiac myocytes rather than neurons. These data shed light on the transcriptional circuits that regulate the differential specification of cardiac myocyte vs. neuronal/glial lineages from a common multipotent progenitor cell. An understanding of this blueprint has implications for our understanding of development and stem cell therapies.
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