Role of TIF1y in Erythropoiesis

Boston Children'S Hospital, Boston MA

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Abstract

Erythropoiesis is driven by an intrinsic transcriptional program that is modified by chromatin factors.[unreadable] Homozygous mutant moonshine mutants are severely anemic and have a complete block in erythroid[unreadable] differentiation at the proerythroblast level. We recently demonstrated that the gene that is defective in the[unreadable] zebrafish moonshine mutant is the transcriptional intermediary factor 1 gamma (TIF1gamma). The TIF1 family has[unreadable] multiple domains including a PhD finger, ring finger and bromo domain, and are thought to bridge DNA[unreadable] binding proteins to other chromatin factors. One published role for TIFs is to modulate transcription[unreadable] regulation by nuclear receptors. TIF1gamma is localized to novel nuclear bodies, and to date no signaling factors[unreadable] have been found to interact with TIF1gamma during erythropoiesis. Recently TIF1gamma has been shown to interact[unreadable] with SMAD factors and regulate epithelial fate during early embryogenesis in frogs, establishing a hypothesis[unreadable] that TGFbeta signaling is abnormal is moonshine mutants. Here we plan to do an extensive characterization of[unreadable] gene expression in moonshine mutants. We plan to sort erythroid progenitors at the onset of the moonshine[unreadable] phenotype, and compare gene expression profiles to wildtype cells, and to other hemtopoietic mutants. The[unreadable] effect of overexpression of TIF1gamma will be evaluated in zebrafish embryos. We plan to use chromatin[unreadable] immunoprecipitation as a method to find targets of TIF1gamma, and will purify TIF1gamma associated proteins using an[unreadable] in vivo biotinylation and streptavidin purification strategy. Particular attention will be given to SMAD factors[unreadable] that might interact with TIF1gamma in erythroid cells at a genetic level. Targeted lesion detection will be used in[unreadable] zebrafish to find an allelic series of moonshine mutants, with mutations in every functional domain of[unreadable] moonshine. Finally, we will undertake a suppressor screen for zebrafish mutants that modify the moonshine[unreadable] phenotype. This should lead to a better understanding of the pathways that moonshine controls, and will[unreadable] have an impact on diseases such as sickle cell anemia and thalassemia.