TRANSCRIPTIONAL REGULATION IN STEM CELLS

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Stage-and tissue specific gene expression controls the ability of hematopoietic stem cells to continue self-renewal versus their proliferation and differentiation into mature, functional hematopoietic cells. This project will focus on the four Id family genes, which encode important negative regulators of the helix-loop-helix (HLH) transcription factor family. Hypothesis; Id family genes are differentially expressed in hematopoietic stem cells during cell cycle progression, cytokine-induced proliferation, and differentiation. Members of this family have been shown to have a profound effect on proliferation and differentiation in a variety of hematopoietic cell lines. Therefore, we propose that members of the Id- family regulate hematopoietic stem cell proliferation and its commitment to differentiation. I. The expression patterns of the four Id family genes will be examined in murine hematopoietic cell lines during self-renewal and induced differentiation. The expression of the four endogenous id family genes will be followed during synchronized growth, and during proliferation and differentiation induced by lineage-selective cytokines. II. The consequences of Id gene overexpression the growth and differentiation of murine hematopoietic cell lines will be tested by transfection of Id gene constructs under transcriptional or selenium- regulated translational control. The ability of Id HLH regulators to affect cell proliferation and differentiation will be assayed using hematopoietic cell lines with integrated, regulated expression constructs of individual Id family genes. III. Id family gene expression will be examined in hematopoietic stem cells and the consequences of overexpression determined on cell cycle progression, differentiation and engraftment into myeloablated mice. Murine stem cells transfected with regulated Id gene constructs will be examined for the effects of overexpression by evaluation of proliferation and differentiation in vitro, as well as on differentiation and engraftment in vivo. The results of these studies will advance our basic understanding of stem cell biology that will, in turn, promote further progress in bone marrow transplantation and gene therapy.