Control of Erythroid Differentiation by Transcription Elongation

项目总结(见说明)： 红系分化由细胞特异性DNA结合蛋白SCL在转录水平上控制 和GATA1。利用斑马鱼的基因组学，我们在红系细胞中发现了一个新的分化检查点，可能 确保了成功的成熟。我们在脊椎动物中进行了第一次隐性抑制基因筛选，以找到一个 在贫血斑马鱼突变体月光的背景下可以恢复红细胞生成的新突变体。 Moonlight编码斑马鱼转录中介因子1伽马(TIFly)的同源基因，以及 月光蛋白的第一个抑制因子编码cdc73，这是一种参与转录延长的染色质因子。 月光突变体暂停了血液特异的rna转录，cdc73缺乏使拉长纤维 发生并挽救红系基因的表达。TIFly与SCL转录因子复合体和PTEFb结合， 一种使PolII磷酸化并调节RNA转录物的伸长的激酶。我们有 检测了条件性TIFly基因敲除小鼠的造血表型。骨髓 红细胞生成缺陷，骨髓生成增加，B细胞严重缺乏。我们计划 评估该突变小鼠红系和髓系祖细胞的转录因子延长。移栽 实验计划评估一种假定的干细胞功能缺陷和红细胞的自主性。 叛逃。我们将确定差异检查点是否在诸如色素减退等疾病中被激活 贫血或膜组装缺陷。HEXIM 1最初被鉴定为在MEL过程中诱导的基因 它与其他蛋白质和7SK RNA结合，调节P-TEFb的活性。我们 推测HEXIM复合体可作为红系分化的传感器。我们计划定义 HEXIM在红系细胞中的结合伙伴，以及HEXIM复合体亚单位的功能 在鱼和老鼠的红细胞生成过程中。HEXIM的7SK RNA组件的功能将是 被投资了。一项新的测序技术将确定是否有其他小RNA与HEXIM结合 很复杂。转录延长在红系细胞分化检查点的调节 与地中海贫血、镰状细胞性贫血、铁缺乏和膜缺陷等疾病有关。

PROJECT SUMMARY (See instructions): Erythroid differenfiafion is controlled at the transcripfional level by the cell-specific DNA-binding proteins SCL and GATA1. Using zebrafish genefics, we found a new differentiation checkpoint in erythroid cells that likely ensures successful maturafion. We undertook the flrst recessive suppressor screen in vertebrates to find a novel mutant that can recover erythropoiesis in the background ofthe anemic zebrafish mutant moonshine. Moonshine encodes the zebrafish ortholog of transcripfional intermediary factor 1 gamma (TIFly), and the first suppressor of moonshine encoded cdc73, a chromatin factor involved in transcripfion elongafion. Moonshine mutants have paused blood-specific RNA transcripts, and cdc73 deficiency allows elongafion to occur and rescues erythroid gene expression. TIFly binds to the SCL transcription factor complex and PTEFb, the kinase that phosphorylates pol II and regulates the elongation of RNA transcripts. We have examined the hematopoiefic phenotype of a condifional mouse knockout for TIFly. Bone marrow erythropoiesis is defective, myelopoiesis is increased, and there is a severe deficiency of B cells. We plan to evaluate transcripfion elongation in erythroid and myeloid progenitors of this mouse mutant. Transplantafion experiments are planned to evaluate a putative stem cell funcfional defect and the autonomy ofthe red cell defect. We will establish if the differenfiafion checkpoint is acfivated in diseases such as hypochromic anemias or membrane assembly defects. HEXIM 1 was originally identified as a gene induced during MEL differenfiation, and it complexes with other proteins and 7SK RNA to regulate P-TEFb activity. We hypothesize that HEXIM complexes could serve as a sensor of erythroid differenfiafion. We plan to define the binding partners of HEXIM in erythroid cells, and to evaluate the funcfion ofthe HEXIM complex subunits during erythropoiesis in fish and mice. The function ofthe 7SK RNA component of HEXIM will be invesfigated. A new sequencing technology will determine if other small RNAs are bound to the HEXIM complex. The regulation of transcription elongation in the differentiation checkpoint of erythroid cells wili be relevant to diseases such as thalassemia, sickle cell anemia, iron deflciency, and membrane defects.