Predoctoral Fellowship for Minority Students

造血是祖细胞分化成不同血细胞谱系的发育过程。Ets蛋白对这一过程至关重要。Ets蛋白PU.1在B细胞、单核细胞、粒细胞、巨核细胞、肥大细胞和未成熟红细胞中表达。与PU.1高度相关的Spi-B主要在B细胞中表达。PU.1和Spi-B结合相同的DNA元素，因此它们可能激活相似的靶基因。PU.1-/-突变小鼠在胚胎期16.5死亡，缺乏B细胞、T细胞、单核细胞和粒细胞。Spi-B /-小鼠存活，但在B细胞中表现出功能缺陷。鉴于突变小鼠表现出不同的表型，本提案中描述的实验旨在确定这些相关蛋白的重叠和独特功能。Spi-B和Ets-1（一种与PU.1高度分化的Ets蛋白）将被插入ES细胞的PU.1位点。为了确定Spi-B和Ets-1是否补足了PU.1缺陷，这些蛋白将通过体外分化观察骨髓细胞进行分析。接下来，我们将使用体外B细胞培养系统和RAG嵌合体小鼠来观察Spi-B对B细胞的拯救作用。最后，产生具有PU.1位点Spi-B的小鼠，以确定胚胎致死性是否被挽救，以及是否产生适当数量的血细胞。上述实验将启发我们了解PU.1和Spi-B共享的功能以及每种蛋白质特有的功能。

Hematopoiesis is the developmental process in which progenitor cells differentiate into different blood cell lineages. Ets proteins are critical to this process. The Ets protein PU.1 is expressed in B cells, monocytes, granulocytes, megakaryocytes, mast cells and immature erythrocytes. Spi-B, which is highly related to PU.1 is predominantly expressed in B cells. PU.1 and Spi-B bind the same DNA elements, so they may activate similar target genes. PU.1-/- mutant mice die at embryonic day 16.5 lacking B cells , T cells, monocytes and granulocytes. Spi-B /- mice are viable but exhibit functional defects in B cells. Given the distinct phenotypes exhibited by mutant mice, the experiments described in this proposal are designed to identify overlapping and unique functions of these related proteins. Spi-B and Ets-1 (an Ets protein highly divergent from PU.1) will be Inserted into the PU.1 locus in ES cells. To determine if Spi-B and Ets-1 complement PU.1 deficiency, these proteins will be analyzed by in vitro differentiation to look at myeloid cells. Next an in vitro B cell culture system and RAG chimera mice will be used to look at B cell rescue by Spi-B. Finally, mice with Spi-B in the PU.1 locus will be generated to determine if the embryonic lethality is rescued and if blood cells are produced in appropriate numbers. The experiments outlined above will enlighten us to the functions PU.1 and Spi-B share and the functions unique to each protein.