Molecular Biology of the alpha IIb/betaPlatelet Receptor

β IIb/β 3整联蛋白受体仅在巨核细胞和血小板表面高度表达。该受体通过纤维蛋白原和其他配体在血小板聚集中起关键作用。拟议的研究将检查巨核细胞生成过程中alphaIIb/beta3的调节表达和alphaIIb/beta3受体的功能生物学。这些研究涉及三个具体目标：一。 定义调节α IIb表达的近端启动子元件，α IIb/β 3的限制性组织表达依赖于α IIb链，因为β 3由许多组织表达。我们已经发现，α IIb表达的关键是造血转录因子加塔-1、FOG，1和Fti-1的复合物与其近端启动子的结合。事实上，许多其他巨核细胞特异性基因的表达调控可能涉及这种相同的复合物的结合。 我们将描述这种复合物的DNA结合位点，以及这种复合物内外与其他核因子相互作用的位点。还将研究与转录因子bKLF结合的alphaIIb近端启动子中的沉默元件。 二.我们已经分析了α IIb基因位点周围>200 kb的区域，并确定了两个遗传上保守的区域，一个对拷贝数依赖性表达重要的上游结构域和一个使表达增强10/3倍的下游结构域。我们计划在体外和转基因小鼠中检查这些区域，以了解这些区域如何分别作为绝缘子和增强子。 三.α IIb/β 3受体的功能分析。我们已经促成了对α IIb的N-末端区域对于配体结合是重要的理解。新的研究表明，该区域也可能定义了阻断纤维蛋白原结合的RGD寡肽敏感性的alphaIIb/beta3物种差异。我们还将研究alphaIIb与beta3的异源二聚化的物种差异。总之，拟议的alphaIIb基因调控的研究将提供新的见解alphaIIb和其他巨核细胞特异性基因是如何在巨核细胞的调控。对α IIb/β 3受体的功能研究将为α IIb/β 3如何结合其配体以及α IIb和β 3链如何异源二聚化提供额外的见解。

The alphIIb/beta3 integrin receptor is highly expressed only on the surface of megakaryocytes and platelets. This receptor has a critical role in platelet aggregation through fibrinogen and other ligands. The proposed studies will examine both the regulated expression of alphaIIb/beta3 during megakaryopoiesis and the functional biology of the alphaIIb/beta3 receptor. These studies involve three specific aims: I. Define the proximal promoter elements regulating alphalIb expression, alphaIIb/beta3's restricted tissue expression is dependent on the alphaIlb chain, since beta3 is expressed by many tissues. We have found that critical to alphaIIb expression is the binding of a complex of hematopoietic transcription factors, GATA-1, FOG,1 and Fti-1, to its proximal promoter. Indeed, the regulated expression of many other megakaryocyte-specific genes may involve binding of this same complex. We will characterize this complex's DNA-binding site, and the sites of interactions within and outside of this complex with other nuclear factors. A silencer element in the alphaIIb proximal promoter that binds the transcription factor bKLF will also be studied. II. Characterize distal regulatory regions of the alphaIIb gene, We have analyzed >200 kb around the alphaIIb gene locus and have defined two phylogenetically-conserved regions, an upstream domain important for copy-number dependent expression and a downstream domain that enhances expression 10/3-fold. We plan to examine these regions both in vitro and in transgenic mice to understand how these regions behave as insulator and enhancer, respectively. III. Functional analysis of the alphaIIb/beta3 receptor. We have contributed to the understanding that an N-terminal region of alphaIIb is important for ligand-binding. New studies show that this region may also define alphaIIb/beta3 species differences in RGD oligopeptide sensitivity for blocking fibrinogen binding. We will also examine species differences in the heterodimerization of alphaIIb with beta3. In summary, the proposed alphaIIb gene regulation studies will provide new insights into how the alphaIIb and other megakaryocyte-specific genes are regulated during megakaryopoiesis. The functional studies on the alphaIlb/beta3 receptor will provide additional insights into how alphaIIb/beta3 bind its ligands, and how the alphaIIb and beta3 chains heterodimerize.