Role of TIF1y in Erythropoiesis

红细胞生成是由一种内在的转录程序驱动的，该程序被染色质因子改变。 纯合突变的月光突变体患有严重贫血，并完全阻断了红系 原红细胞水平的分化。我们最近证明了在人类基因组中存在缺陷的基因 斑马鱼月光突变体是转录中介因子1伽马(TIF1伽马)。TIF1家族拥有 多个结构域，包括PHD手指、无名指和溴结构域，被认为是连接DNA的桥梁 结合蛋白质与其他染色质因子。已发表的TIF的一个作用是调节转录 核受体的调节作用。TIF1Gamma定位于新的核体，到目前为止还没有信号因子 已被发现在红细胞生成过程中与TIF1伽马相互作用。最近，TIF1伽马被证明可以相互作用 与SMAD因子一起调控青蛙早期胚胎发生中的上皮命运，建立了一个假说 转化生长因子β信号异常是兼职突变体。在这里，我们计划做一个广泛的描述 月光突变体中的基因表达。我们计划在月光开始时对红系祖细胞进行分类 表型，并将基因表达谱与野生型细胞和其他造血突变体进行比较。这个 在斑马鱼胚胎中过表达TIF1伽马的效果将被评估。我们计划使用染色质 免疫沉淀作为寻找TIF1-γ靶点的一种方法，并将利用 体内生物素化和链霉亲和素纯化策略。将特别关注SMAD因素 这可能与红系细胞中的TIF1伽马在基因水平上相互作用。靶向病变检测将用于 斑马鱼寻找等位基因系列的兼职突变体，在每个功能区的突变 月光酒。最后，我们将对修饰月光的斑马鱼突变体进行抑制筛选 表型。这应该会导致更好地理解月光控制的途径，并将 对镰状细胞性贫血和地中海贫血等疾病有影响。

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