A platelet integrin, allbbeta3, plays critical roles in platelet adhesion and aggregation. Integrin allbbeta3 mediates two-way signaling transduction. Intracellular signals, via the integrin cytoplasmic domains induces inside-out signaling, activating the ligand binding function of allbbeta3. Ligand binding to allbbeta3 induces outside-in signaling, which also requires integrin cytoplasmic domains to mediate cellular responses such as protein tyrosine phosphorylation, cytoskeleton reorganization, cell spreading, and stable adhesion. We have reconstituted beta3 integrin signaling in cultured mammalian cell model expressing recombinant human integrin llbbeta3 and human glycoprotein Ib-IX. Using this model, we have provided evidence that the structural requirements for outside-in signaling are different from inside-out signaling. Thus we hypothesize that outside-in signaling and inside-out signaling of beta3 integrins can be differentially regulated. Furthermore, we have shown that the cytoplasmic domain of integrin can be cleaved by the calcium-dependent protease, calpain, at specific sites flanking two functionally important NXXY motifs, causing differential regulation of integrin two-way signaling. Thus we further hypothesize that calpain cleavages serve as an important mechanism that differentially regulates inside-out and outside-in signaling. In particular, calpain cleavage at Y759 selectively regulates outside-in signaling. To test these hypotheses, we propose to investigate (1) why common and distinct structures in the cytoplasmic domain of beta3 are required in inside-out and outside-in integrin signaling, (2) the structural basis of calpain recognition of beta3, (3) whether calpain cleavage is regulated by tyrosine phosphorylation, and (4) whether and how calpain regulate integrin signaling using calpain I knockout mice and siRNA techniques and a calpain-resistant mutant the integrin.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL080264-04
Application #
7747903
Study Section
Special Emphasis Panel (ZRG1-HEME-C (02))
Program Officer
Kindzelski, Andrei L
Project Start
2006-12-15
Project End
2011-07-31
Budget Start
2009-12-01
Budget End
2011-07-31
Support Year
4
Fiscal Year
2010
Total Cost
$348,750
Indirect Cost
Name
University of Illinois at Chicago
Department
Pharmacology
Type
Schools of Medicine
DUNS #
098987217
City
Chicago
State
IL
Country
United States
Zip Code
60612
Estevez, Brian; Du, Xiaoping (2017) New Concepts and Mechanisms of Platelet Activation Signaling. Physiology (Bethesda) 32:162-177
Delaney, M Keegan; Kim, Kyungho; Estevez, Brian et al. (2016) Differential Roles of the NADPH-Oxidase 1 and 2 in Platelet Activation and Thrombosis. Arterioscler Thromb Vasc Biol 36:846-54
Estevez, Brian; Kim, Kyungho; Delaney, M Keegan et al. (2016) Signaling-mediated cooperativity between glycoprotein Ib-IX and protease-activated receptors in thrombin-induced platelet activation. Blood 127:626-36
Ju, Lining; Chen, Yunfeng; Xue, Lingzhou et al. (2016) Cooperative unfolding of distinctive mechanoreceptor domains transduces force into signals. Elife 5:
Estevez, Brian; Shen, Bo; Du, Xiaoping (2015) Targeting integrin and integrin signaling in treating thrombosis. Arterioscler Thromb Vasc Biol 35:24-9
Shen, Bo; Estevez, Brian; Xu, Zheng et al. (2015) The interaction of G?13 with integrin ?1 mediates cell migration by dynamic regulation of RhoA. Mol Biol Cell 26:3658-70
Delaney, Michael Keegan; Liu, Junling; Kim, Kyungho et al. (2014) Agonist-induced platelet procoagulant activity requires shear and a Rac1-dependent signaling mechanism. Blood 124:1957-67
Rusu, Luiza; Andreeva, Alexandra; Visintine, David J et al. (2014) G protein-dependent basal and evoked endothelial cell vWF secretion. Blood 123:442-50
Li, Jing; Kim, Kyungho; Hahm, Eunsil et al. (2014) Neutrophil AKT2 regulates heterotypic cell-cell interactions during vascular inflammation. J Clin Invest 124:1483-96
Du, Xiaoping (2014) Self-control of platelets: a new ITIM story. Blood 124:2322-3

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