Molecular Biology of the alpha IIb/betaPlatelet Receptor

University Of Pennsylvania, Philadelphia PA

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Abstract

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.