Modulation of Leukocyte Adhesion by P-selectin/PSGL-1 Signaling

描述(由申请人提供)：P-选择素/PSGL-1信号调节白细胞黏附需要一系列细胞事件，包括反应细胞的滚动和牢固黏附。P-选择素(CD62P)、P-选择素糖蛋白配体-1(PSGL-1、CD162)和β2-整合素是天然免疫和炎症所必需的内皮细胞和白细胞黏附分子。P-选择素与PSGL-1的相互作用介导了白细胞的滚动，在此过程中，它们被局部释放或展示的细胞因子和趋化物质充分激活，以促进β2整合素介导的牢固黏附。然而，P-选择素黏附活性的反馈调节机制和P-选择素诱导的β2-整合素活化机制尚不清楚。NEF相关因子1(NAF1)是核因子-kappaB激活的内源性抑制因子。血清淀粉样蛋白P组分(SAP)是肝细胞在各种炎症介质作用下快速合成和分泌的一种急性期蛋白。在初步研究中，我们发现NaF1与PSGL-1的胞质尾部形成了稳定的复合体。P-选择素激活PSGL-1使NAF1的Y552PPM基序磷酸化，以招募磷脂酰肌醇-3激酶(PI3K)的P85亚单位，并触发信号级联，最终导致αMbeta2(CD11bCD18，Mac-1)的激活。此外，我们还观察到SAP与P-选择素相互作用，并作为PSGL-1的内源性抑制物识别P-选择素。在这项拨款申请中，我们建议1)确定P-选择素诱导的αMbeta2激活的功能重要性；2)确定PSGL-1-NAF1-P85信号通路对调节αMbeta2活性的生物学意义；以及3)探讨SAP对P-选择素黏附活性的调节。总体而言，这项研究不仅将加深我们对在白细胞募集的多步骤范式中精确决定白细胞命运的分子机制的理解，而且还将发现预防和治疗炎症性疾病的新的治疗靶点。

DESCRIPTION (provided by applicant): Modulation of Leukocyte Adhesion by P-selectin/PSGL-1 Signaling Leukocyte recruitment entails a cascade of cellular events, including rolling and firm adhesion of responding cells. P-selectin (CD62P), P-selectin glycoprotein ligand-1 (PSGL-1, CD162) and beta2-integrins are endothelial and leukocyte cell adhesion molecules essential for innate immunity and inflammation. The interaction of P-selectin with PSGL-1 mediates leukocyte rolling, during which they become sufficiently activated in situ by locally released or displayed cytokines and chemoattractants for beta2-integrin-mediated firm adhesion. However, the mechanisms for feedback regulation of P-selectin adhesion activity and P- selectin-induced beta2-integrin activation remain undetermined. Nef-associated factor 1 (Nafl) is an endogenous inhibitor for NF-kappaB activation. Serum amyloid P component (SAP) is an acute phase protein synthesized and secreted rapidly by hepatocytes in response to various inflammatory mediators. In the preliminary studies, we found that Naf1 formed a stable complex with the cytoplasmic tail of PSGL-1. Engagement of PSGL-1 by P-selectin phosphorylated the Y552PPM motif of Naf1 for recruiting p85 subunit of phosphatidylinositol-3 kinase (PI3K) and triggering the signaling cascade that culminated in activation of alphaMbeta2 (CD11bCD18, Mac-1). In addition, we observed that SAP interacted with P-selectin and acted as an endogenous inhibitor of PSGL-1 for P-selectin recognition. In this grant application, we propose 1) to define the functional importance of P-selectin-induced activation of alphaMbeta2; 2) to determine the biological significance of the PSGL-1-Naf1-p85 signaling pathway for modulation of alphaMbeta2 activity; and 3) to explore the regulation of P-selectin adhesion activity by SAP. Overall, this study will not only enhance our understanding of molecular mechanisms that precisely determine leukocyte fates in the multi- step paradigm of leukocyte recruitment, but also discover novel therapeutic targets for prevention and treatment of inflammatory disorders.