Interactions with P- and E-selectin cause flowing neutrophils to roll on activated platelets and endothelial cells. Rolling neutrophils activate b2 integrins that interact with intercellular adhesion molecule-1 (ICAM-1) to slow rolling, induce arrest, and enable emigration into surrounding tissues. Neutrophil adhesion and signaling are interdependent. In the past funding period we explored mechanisms for selectin-triggered, partial aLb2 activation to slow rolling and for chemokine-triggered, full aLb2 activation to mediate arrest. We discovered that chemokines also induce aLb2-dependent slow rolling. We distinguished talin-docking requirements for partial and full activation of aLb2. We adopted a system to express WT or mutant proteins in immortalized myeloid progenitors that are differentiated into neutrophils to probe their functions. In the next grant cycle we will further explore the molecular basis for signal-dependent neutrophil adhesion under flow. We will use biochemical assays, flow-chamber experiments, and intravital microscopy to distinguish components used for inside-out activation of aLb2 by selectins or chemokine and for outside-in signaling by integrins. To define how the cytoplasmic domain of P-selectin glycoprotein ligand-1 (PSGL-1) initiates signaling, we will generate PSGL-1 cytoplasmic-domain mutants in differentiated neutrophils from PSGL-1-deficient mice. We will screen for mutants that do not activate kinases or trigger aLb2-dependent slow rolling. We will use mass spectrometry to Identify cytosolic proteins that bind to the WT PSGL-1 tail but not to signal-defective tail mutants. We will define steps in talin1-dependent b2 integrin activation by expressing WT or mutant talin1 or b2 in neutrophils differentiated from talin1- or b2-deficient myeloid progenitors. These studies will yield new insights into connections between neutrophil adhesion and signaling that may offer improved methods to treat inflammatory and thrombotic diseases.
White blood cells (leukocytes) combat infection and repair tissue injury. To do this, they must move from blood into the affected tissues, a process that begins when circulating leukocytes attach to blood vessel surfaces through adhesion receptors. This project addresses how leukocytes and cells lining blood vessels control the display and function of these receptors. Because excessive recruitment of leukocytes contributes too many diseases, including heart attacks and strokes, understanding how cells regulate adhesion receptors may suggest new therapeutic strategies.
|Sreeramkumar, Vinatha; Adrover, José M; Ballesteros, Ivan et al. (2014) Neutrophils scan for activated platelets to initiate inflammation. Science 346:1234-8|
|Zhang, Yan; Jiang, Ning; Zarnitsyna, Veronika I et al. (2013) P-selectin glycoprotein ligand-1 forms dimeric interactions with E-selectin but monomeric interactions with L-selectin on cell surfaces. PLoS One 8:e57202|
|Zarbock, Alexander; Ley, Klaus; McEver, Rodger P et al. (2011) Leukocyte ligands for endothelial selectins: specialized glycoconjugates that mediate rolling and signaling under flow. Blood 118:6743-51|
|Miner, Jonathan J; Shao, Bojing; Wang, Ying et al. (2011) Cytoplasmic domain of P-selectin glycoprotein ligand-1 facilitates dimerization and export from the endoplasmic reticulum. J Biol Chem 286:9577-86|
|Shao, Bojing; Wahrenbrock, Mark G; Yao, Longbiao et al. (2011) Carcinoma mucins trigger reciprocal activation of platelets and neutrophils in a murine model of Trousseau syndrome. Blood 118:4015-23|
|Yago, Tadayuki; Fu, Jianxin; McDaniel, J Michael et al. (2010) Core 1-derived O-glycans are essential E-selectin ligands on neutrophils. Proc Natl Acad Sci U S A 107:9204-9|
|Yago, Tadayuki; Shao, Bojing; Miner, Jonathan J et al. (2010) E-selectin engages PSGL-1 and CD44 through a common signaling pathway to induce integrin alphaLbeta2-mediated slow leukocyte rolling. Blood 116:485-94|
|McEver, Rodger P; Zhu, Cheng (2010) Rolling cell adhesion. Annu Rev Cell Dev Biol 26:363-96|
|Miner, Jonathan J; Xia, Lijun; Yago, Tadayuki et al. (2008) Separable requirements for cytoplasmic domain of PSGL-1 in leukocyte rolling and signaling under flow. Blood 112:2035-45|
|Yago, Tadayuki; Lou, Jizhong; Wu, Tao et al. (2008) Platelet glycoprotein Ibalpha forms catch bonds with human WT vWF but not with type 2B von Willebrand disease vWF. J Clin Invest 118:3195-207|
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