Leukocyte integrin aMb2 (CD11b/CD18, Mac-1) plays a pivotal role in normal protective inflammatory response and pathological inflammation. This receptor has prodigious adhesive and signaling capabilities which allowed it to become the premier workhorse in host defense. It is also a potential therapeutic target in many diseases in which inflammation plays an essential role, including cardiovascular diseases. The diverse functions and activities ascribed to aMb2 arise from its ability to bind a multitude of structurally diverse ligands. However, the mechanisms which allow aMb2 to exhibit broad ligand recognition are still poorly understood. Our previous studies with a prototype aMb2 ligand fibrinogen provided initial insight into the mechanism by which the aMI-domain of the receptor recognizes its ligands. In the past funding period we have solved the consensus aMI-domain recognition motif, we termed IRM. A key feature of IRM is a small core consisting of specific combinations of basic and hydrophobic amino acid residues ubiquitous in many aMb2 ligands. The characteristics of IRM are consistent with the capacity of aMb2 to recognize a wide variety of unrelated sequences and, thus, form a molecular basis for aMb2 ligand binding promiscuity.
Specific Aim1 is to further characterize the mechanism underlying broad recognition specificity of aMb2. Combinatorial peptide libraries and mutational analyses will be used to clarify the structural features of IRM. Mass spectrometry will be used to determine the effect of inflammation-associated protein modifications on the function of IRM. Our preliminary studies revealed that neutrophil secretion products are enriched in IRMs which allowed their prediction as a new class of aMb2 ligands. We have found that one of them, human neutrophil cathelicidin peptide LL-37, effectively binds aMb2 and unduces a potent aMb2- dependent migratory response. Based on this finding we propose that LL-37 and other neutrophil-derived proteins/peptides exert their potent immunomodulatory effects by binding aMb2 on monocyte/macrophages.
Specific Aim 2 is to test this hypothesis by characterizing aMb2-dependent monocyte responses elicited by LL-37. The effect of LL-37 on signaling and migratory functions of aMb2 will be determined using aMb2- expressing and aMb2-deficient cells and in the in vivo animal model. Studies over the past funding period identified integrin aDb2 as a multiligand receptor with specificity similar to that of aMb2 and revealed that its upregulation on inflammatory macrophages inhibits their migration.
Specific Aim 3 is to characterize the role of aMb2 and aDb2, two most abundant and adhesive integrins on macrophages, in emigration of these cells from the inflammatory site during the resolution of inflammation. The efflux of macrophages by draining lymphatics will be investigated in wild-type and integrin-deficient mice. Overall, these studies will lead to an increased understanding of the principles which govern ligand recognition by aMb2, will give new insights into the biology of aMb2 and aDb2 and may be useful in the design of novel therapeutic strategies.

Public Health Relevance

Inflammation is critically involved in the pathogenesis of many disorders, including cardiovascular disease. Integrin aMb2 (Mac-1) is the most versatile receptor on leukocytes and mediates numerous responses of these cells during the inflammatory response. The multiplicity of functions exhibited by Mac-1 depends on its ability to bind a myriad of diverse proteins. Understanding the molecular basis for the extreme stickiness of Mac-1 and the biological significance of receptor's broad recognition could lead to new methods of treatment of disorders in which inflammation plays a role.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL063199-14
Application #
8585079
Study Section
Hemostasis and Thrombosis Study Section (HT)
Program Officer
Kindzelski, Andrei L
Project Start
1999-07-01
Project End
2014-11-30
Budget Start
2013-12-01
Budget End
2014-11-30
Support Year
14
Fiscal Year
2014
Total Cost
$343,125
Indirect Cost
$118,125
Name
Arizona State University-Tempe Campus
Department
Other Basic Sciences
Type
Schools of Arts and Sciences
DUNS #
943360412
City
Tempe
State
AZ
Country
United States
Zip Code
85287
Podolnikova, Nataly P; Yakovlev, Sergiy; Yakubenko, Valentin P et al. (2014) The interaction of integrin *IIb*3 with fibrin occurs through multiple binding sites in the *IIb *-propeller domain. J Biol Chem 289:2371-83
Lishko, V K; Yermolenko, I S; Owaynat, H et al. (2012) Fibrinogen counteracts the antiadhesive effect of fibrin-bound plasminogen by preventing its activation by adherent U937 monocytic cells. J Thromb Haemost 10:1081-90
Yermolenko, Ivan S; Fuhrmann, Alexander; Magonov, Sergei N et al. (2010) Origin of the Nonadhesive Properties of Fibrinogen Matrices Probed by Force Spectroscopy. Langmuir :
Podolnikova, Nataly P; O'Toole, Timothy E; Haas, Thomas A et al. (2009) Adhesion-induced unclasping of cytoplasmic tails of integrin alpha(IIb)beta3. Biochemistry 48:617-29
Yakubenko, Valentin P; Belevych, Nataly; Mishchuk, Daria et al. (2008) The role of integrin alpha D beta2 (CD11d/CD18) in monocyte/macrophage migration. Exp Cell Res 314:2569-78
Yakubenko, Valentin P; Yadav, Satya P; Ugarova, Tatiana P (2006) Integrin alphaDbeta2, an adhesion receptor up-regulated on macrophage foam cells, exhibits multiligand-binding properties. Blood 107:1643-50
Yakovlev, Sergiy; Zhang, Li; Ugarova, Tatiana et al. (2005) Interaction of fibrin(ogen) with leukocyte receptor alpha M beta 2 (Mac-1): further characterization and identification of a novel binding region within the central domain of the fibrinogen gamma-module. Biochemistry 44:617-26
Podolnikova, Nataly P; Gorkun, Oleg V; Loreth, Ralph M et al. (2005) A cluster of basic amino acid residues in the gamma370-381 sequence of fibrinogen comprises a binding site for platelet integrin alpha(IIb)beta3 (glycoprotein IIb/IIIa). Biochemistry 44:16920-30
Lishko, Valeryi K; Podolnikova, Nataly P; Yakubenko, Valentin P et al. (2004) Multiple binding sites in fibrinogen for integrin alphaMbeta2 (Mac-1). J Biol Chem 279:44897-906
Schober, Joseph M; Lau, Lester F; Ugarova, Tatiana P et al. (2003) Identification of a novel integrin alphaMbeta2 binding site in CCN1 (CYR61), a matricellular protein expressed in healing wounds and atherosclerotic lesions. J Biol Chem 278:25808-15

Showing the most recent 10 out of 20 publications