Regulation of Integrin Signaling by Adapter Proteins

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Integrins are dimeric cell surface receptors that play critical roles in cellular adhesion and migration. Signaling events critical for integrin function include second messenger cascades initiated by other cell surface receptors which modulate integrin avidity for their ligands in addition to signaling pathways initiated by engagement of integrins themselves. We and others have found that the formation of multimolecular complexes nucleated by adapater proteins is essential for both signaling to integrins (inside-out signaling) and signaling by integrins (outside-in signaling). Our laboratory has a long standing interest in the SH2 domain containing leukocyte phosphoprotein of 76kDa (SLP-76), a hematopoietic restricted adapater protein that is critical for function of both immunoreceptors and integrins. The experiments described in this proposal will make use of biochemical, imaging, molecular and genetic approaches to investigate the role of SLP-76 and its associated molecules in integrin responses in cells of both the innate and adaptive immune systems. The work will be divided into three specific aims. The first will explore the inducible interactions between SLP-76 and other proteins upon integrin engagement and the subcellular localization of SLP-76 during integrin function in T cells. One central hypothesis that will be tested is that there are two pools of SLP-76 in the cell, one which regulates immunoreceptor signaling and the second which regulates integrin responses. We will further test the notion that integrin pathway requires an interaction between SLP-76 and three adapter proteins, ADAP (Adhesion and Degranulation-promoting Adapter Protein), SKAP55 (SrcKinase- associated phosphoprotein of 55kDa), and RIAM (Rap1-GTP interacting adapter molecule), and that these adapters collectively nucleate a complex which brings activated Rap-1 to the cell surface. The second aim of this proposal will take the biochemical and imaging studies of Aim 1 in vivo by generating genetically unique murine lines in which mutations of SLP-76 or its associated molecules are expressed that are predicted to selectively impact T cell immunoreceptor or integrin function. The third aim will extend our work to the innate immune system by studying the biochemistry and molecular events coordinated by these adapter molecules in neutrophils, both ex vivo and in vivo. Experiments described in this project will make extensive use of both scientific cores and will rely extensively on interactions with the Project Leaders of the other projects of this program. We hope that these studies will provide new insights into how multimolecular complexes integrate signaling pathways leading to appropriate immune cell responses.