Project summary/Abstract. Long-term hematopoietic stem cells (HSCs) are capable of self-renewal and differentiation into all mature hematopoietic lineages. The established paradigm for this process involves the self-renewal and differentiation of HSCs into progenitor cells. The primary goal of our proposal is to define mechanisms to enhance self-renewal in stem and progenitor cells for hematopoietic stem cell transplantation in patients. A steady rise in patients receiving HSC transplants for non-malignant diseases and disorders has occurred over the last two decades, but survival of these patients can be as low as 20-40% in the five year post-transplant period. This suggests a strong need for improved HSC transplants for these patients. To advance our understanding of HSC function and self-renewal, I completed the first large-scale in vivo reverse genetic screen targeting chromatin factors, which regulate gene expression through changes in chromatin structure. From this screen, we found a requirement for the plant homeodoming (PHD) finger and chromatin associated protein, Ing4. As a member of the Hbo1 chromatin remodeling complex, Ing4 binds to target loci through recognition of H3K4me3 marks and increases localized levels of histone acetylation. Ing4 has also been shown to negatively regulate NF-κB, c-Myc, Hif1a, and p53. Our work in Ing4-deficient mice has shown that loss of Ing4 has a dramatic effect on hematopoiesis in the bone marrow. These mice show increased levels of HSCs and a loss of multipotent progenitors (MPPs). Transplantation of MPPs shows that while there are fewer MPPs, the cells that do differentiate to the MPP stage are more capable of robust, long-term reconstitution than their wild-type counterparts, showing very high levels of chimerism up to 9 months post- transplant. We hypothesize that Ing4 regulates self-renewal properties in MPP cells and loss of Ing4 results in MPPs with activated stem cell-like properties. Work done under this proposal will identify the pathways altered in Ing4-deficient MPPs to characterize how they have regained the ability to function like HSCs. If successful, our studies will provide a new mechanism for re-activating self-renewal pathways. This could lead to more successful HSC transplants for patients needing this treatment.

Project Narrative The molecular mechanisms that regulate stem cell self-renewal are not well characterized. The studies outlined in this proposal aim to define how Ing4 deficiency results in the reprogramming of a progenitor cell population that normally would not be capable of self-renewal. Our work will broaden our understanding of the components that regulate self-renewal in hematopoietic stem cell fate decisions and may help identify therapeutic targets that could be useful in the treatment of hematological diseases where robust stem cell activity is needed for stem cell transplantation.