Over the last 2 years array and mass spectrometry technologies have enabled analysis of the transcriptome and proteome of hematopoietic stem cells (HSC) and their downstream progenitors. This information will be of significant value to the elucidation of molecular mechanisms that govern stem cell physiology and blood cell differentiation. However, an equally, if not more important goal, is to define those proteins that participate in signaling pathways that are active in hematopoietic stem cells and their progeny. Enzymes that phosphorylate tyrosine, serine and threonine residues on other proteins play a major role in signaling cascades that determine cell cycle entry, survival and the differentiation fate of cells in the mammalian body, including the hematopoietic system. In particular, knowing the signaling pathways that are active in HSC and their differentiated progeny will provide critical information for understanding stem cell biology and cell differentiation. Traditional genetic and biochemical approaches can certainly provide some of these answers; however, for technical and practical reasons, these are typically pursued one gene or pathway at a time. Thus, a more comprehensive approach is needed in order to reveal signaling pathways active in HSC and their differentiated progeny. Towards this end, we have developed kinome analysis techniques and begun to apply them to HSC and differentiated progenitor cell types. In this study, we propose to provide a comprehensive analysis of the HSC kinome and their more differentiated progeny as this will further define our understanding of the signaling pathways utilized during different stages of hematopoietic development. This proposal will be pursued in the following specific aims: Aim 1: Compare and contrast the kinome of LT-HSC and ST-HSC populations during steady-state hematopoiesis. Aim 2: Determine the LT-HSC kinome during expansion of the hematopoietic compartment. Aim 3: Determine the kinome of key multi-potent progenitors and lineage-restricted progenitor cell types.