Hematopoietic stem cells (HSCs) are the functional units of bone marrow transplantation, which is used in the treatment of variety of blood cell diseases including leukemia and autoimmunedisorders. Clinical use of HSCs is limited by the fact that they are rare cells, occurring at afrequency of only <1/20,000 bone marrow cells. Efforts to expand HSCs prior to transplant by exvivo culturing have proven challenging and thus far such efforts have not translated to the clinic.Thus there remains a clinical need to find alternative strategies for either expanding thenumbers of HSCs, or generating HSCs de novo. Numerous studies have shown that it ispossible to experimentally reprogram the cellular identity of one cell type to another by enforcedexpression of transcription factors involved in the specification of the target cell type. The long-term objective of this project is to use this experimental paradigm and express transcriptionfactors involved in specification of HSC fate and function in committed blood cells to reprogramthem back to an induced stem cell fate. In order to achieve this we must first identify the factorsinvolved in specifying the fate and function of HSCs. This will be achieved using a large-scaleexpression profiling strategy designed to allow us to identify transcription factors uniquelyexpressed in HSCs compared to the other cells of the blood system. Our preliminary studieshave identified 28 such genes. Once identified we will then clone these factors into an induciblelentiviral delivery system to allow us to combinatorially express these genes in committed bloodcells. We will then test whether or not committed blood cells receiving such factors are able offunction like HSCs using a wide variety of stem cell assays. Once reprogramming has beenachieved, it will then be important to determine the minimal combination of factors capable of mediating reprogramming. We will further examine the fidelity of reprogramming by functional,molecular and epigenetic analyses. Successful identification of factors capable ofreprogramming committed blood cells to an induced HSC fate has the potential to advance ourbasic understanding of hematopoietic stem cell biology, and contribute to their clinical utility in anumber of ways. For example, identification of factors capable of reprogramming committedblood cells to HSCs could provide important insights the molecular mechanisms involved inspecifying the fundamental stem cell properties of self-renewal and multi-potency. In terms of clinical application, if successful, our study opens the possibility of eventually generating patient-specific induced HSCs for use in bone marrow transplantation, or for the study of blooddisorders.