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.

Public Health Relevance

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.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
2R01HL034363-29
Application #
8693189
Study Section
Intercellular Interactions (ICI)
Program Officer
Sarkar, Rita
Project Start
1987-09-01
Project End
2018-06-30
Budget Start
2014-09-03
Budget End
2015-06-30
Support Year
29
Fiscal Year
2014
Total Cost
$427,500
Indirect Cost
$177,500
Name
Oklahoma Medical Research Foundation
Department
Type
DUNS #
077333797
City
Oklahoma City
State
OK
Country
United States
Zip Code
73104
Liu, Zhenghui; Zhang, Nan; Shao, Bojing et al. (2016) Replacing the Promoter of the Murine Gene Encoding P-selectin with the Human Promoter Confers Human-like Basal and Inducible Expression in Mice. J Biol Chem 291:1441-7
Zhang, Nan; Liu, Zhenghui; Yao, Longbiao et al. (2016) P-Selectin Expressed by a Human SELP Transgene Is Atherogenic in Apolipoprotein E-Deficient Mice. Arterioscler Thromb Vasc Biol 36:1114-21
Shao, Bojing; Yago, Tadayuki; Setiadi, Hendra et al. (2015) O-glycans direct selectin ligands to lipid rafts on leukocytes. Proc Natl Acad Sci U S A 112:8661-6
Pruenster, Monika; Kurz, Angela R M; Chung, Kyoung-Jin et al. (2015) Extracellular MRP8/14 is a regulator of β2 integrin-dependent neutrophil slow rolling and adhesion. Nat Commun 6:6915
Yago, Tadayuki; Tsukamoto, Hiroki; Liu, Zhenghui et al. (2015) Multi-Inhibitory Effects of A2A Adenosine Receptor Signaling on Neutrophil Adhesion Under Flow. J Immunol 195:3880-9
McEver, Rodger P (2015) Selectins: initiators of leucocyte adhesion and signalling at the vascular wall. Cardiovasc Res 107:331-9
Yago, Tadayuki; Petrich, Brian G; Zhang, Nan et al. (2015) Blocking neutrophil integrin activation prevents ischemia-reperfusion injury. J Exp Med 212:1267-81
Wang, Ying; Yago, Tadayuki; Zhang, Nan et al. (2014) Cytoskeletal regulation of CD44 membrane organization and interactions with E-selectin. J Biol Chem 289:35159-71
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

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