We have discovered that quantitative expression of the stem cell expressed transcriptional represser Transforming-growth-interfering factor (TGIF) is the most discriminating and powerful predictor of patient survival identified in acute myelogenous leukemia (AML). Lower levels of TGIF correlated with shorter survival, whereas higher levels predicted longer survival (p=0.00001). To understand the biological basis for our clinical finding, we investigated TGIF's function in myeloid progenitor cell lines and in a Tg/'f knockout mouse model. TGIF knockdown in myeloid progenitor cell lines, such as HL60, resulted in significant growth inhibition, a relative differentiation block, and disruption of the myeloid transcription program. Analysis of hematopoietic stem cell (HSC) function in Tg/7-null mice showed that the HSCs produced fewer colony- forming units and had a growth advantage compared to wild-type HSCs in competitive transplant studies. These data suggest that Tg/f knockout may alter the balance between the key HSC functions of quiescence, differentiation and self-renewal. We thus hypothesize that Tgif plays a fundamental regulatory role in HSC function and will examine this hypothesis by 1) defining more thoroughly the role of Tgif'm HSC and progenitor cell function;2) determining the cellular basis of Tgif's effects on HSCs;and 3) determining the molecular basis of Tgif's effects on HSCs.
These aims will be carried out in a series of experiments using a variety of standard and state-of-the-art molecular techniques, including limiting dilution competitive repopulation analysis, serial transplantation, in vivo cell cycle analysis, and structure-function characterizations using naturally occurring TGIF mutations and ex vivo HSC transduction. In addition to the Tgif knockout model currently in use, we will also employ a new conditional Tgif knockout allele, which will offer us great flexibility in our experimental design. This proposal also outlines a plan designed to help the candidate develop a successful career in academic medicine. The candidate has a Ph.D. in Microbiology and Immunology from Vanderbilt University and has completed residency training in Pediatrics and fellowship training in Genetics. The candidate will expandhis knowledge of hematopoiesis and leukemogenesis under the mentorship of Drs. Stephen Brandt and Scott Hiebert. A mentoring committee will provide added guidance at regular intervals.
In acute leukemias, including AML, the normal balance between quiescence, self-renewal and differentiation of HSCs is disrupted. Our preliminary data suggest that Tgif may regulate this balance and that it also has a clinical role as a prognostic indicator in AML. Thus, our investigations of how Tgif regulates key HSC functions will provide new insights into HSC biology as well as increase our understanding of AML pathogenesis, leading to better diagnosis and treatment of this devastating disease.
