Critical roles of GA binding protein in HSC maintenance and ageing

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DESCRIPTION (provided by applicant): Hematopoietic stem cells (HSCs) have two defining features. One is the ability to self-renew, maintaining a constant number under steady state, and the other is the potential to differentiate into multiple lineages in blood, meeting the regenerative needs. The maintenance of HSC homeostasis and multipotency is stipulated by transcription factors such as Bmi1, Zfx, and Foxo proteins. However, it is unknown if these factors interact with each other and if they form a hierarchical gene regulatory network in HSCs. In this project, we will investigate the roles of GA binding protein (GABP) in regulating HSC homeostasis and repopulation capacity. We will further characterize its interplay with other transcription factors in controlling different aspects of HSC biology. A functional GABP complex contains one DNA binding subunit, GABP1, and one transactivation subunit, GABP2. Two of GABP2 isoforms contribute to the formation of GABP 1222 tetramers. Inactivation of GABP1 abrogates the activity of entire GABP complex, causing early embryonic lethality. In contrast, mice with deletion of both tetramer-forming GABP2 isoforms were viable. Both strains of animals showed diminished HSC frequency and numbers, but differed in onset and kinetics of the phenotypes. We hypothesize that the GABP complex controls different pathways regulating HSC homeostasis and function and the 1222 tetramers regulate a subset of GABP target gene and have a specific role in regulating HSC self- renewal and ageing. Our long-term goal is to elucidate the interplay between GABP and other key transcription factors and how such interplay balances self-renewal and differentiation of HSCs. This project will generate critical information for a better understanding of HSC plasticity, hematopoietic malignancy, and stem cell ageing. We will approach these objectives through the following specific aims: SPECIFIC AIM 1. To investigate the roles of GABP complex in HSC self-renewal and repopulation capacity. We will use a tissue-specific GABP1-targeted mouse strain, induce inactivation of GABP1 in the bone marrow, and determine the impact on HSC self-renewal and repopulation capacity. We will also investigate if HSC cell cycle and survival pathways are affected by GABP1 deficiency. SPECIFIC AIM 2. To identify GABP direct target genes which mediate its regulation of HSC function. We have obtained a genome-wide GABP binding map in HSCs and found direct GABP binding to regulatory regions of a number of transcription factors. We will characterize the interplay between GABP and these transcription factors to construct a gene regulatory network maintaining HSC homeostasis and multipotency. SPECIFIC AIM 3. To determine a specific role of GABP 1222 tetramers in DNA damage repair and HSC ageing. We will use the mouse strain that is deficient for both tetramer-forming GABP2 isoforms and determine the specific aspect of HSC biology that is controlled by GABP 1222 tetramers. We will also investigate the direct link of tetramers to DNA damage repair and HSC ageing. PUBLIC HEALTH RELEVANCE: Hematopoietic stem cells (HSCs) are responsible for sustained production of all blood cells throughout one's life, and HSCs have two defining features, self-renewal to maintain a constant number and plasticity to generate multiple blood lineages. Aberrant regulation of HSCs can either lead to defective regeneration of blood cells or result in hematopoietic malignancy. This project will investigate critical roles of a transcription factor name GA binding protein and characterize the interplay between GA binding protein and other factors in regulating HSC self-renewal and repopulation capacity, and these studies will lead to a better understanding on how HSC homeostasis and multi-potency are controlled, and provide a molecular basis on how to maintain HSCs to meet regenerative needs and prevent malignant transformation.