The beta2 subfamily of integrins, which includes alphaMbeta2, controls the participation of leukocytes in biological settings. A number of specific leukocyte responses are regulated by alphaMbeta2, including the respiratory burst, homotypic aggregation, phagocytosis, migration, apoptosis, tumor surveillance, and ischemia-reperfusion injury. These responses depend upon the capacity of alphaMbeta2, to function as a receptor for an extremely broad spectrum of ligands. The mechanism by which alphaMbeta2 interacts with structurally diverse ligands to mediate the cellular responses of adhesion, migration and activation is the central focus of this application. Preliminary data developed in this application indicate that four representative ligands, iC3b, NIF, the P2 peptide of fibrinogen and C. albicans, not only interact with the I-domain in the alphaM subunit but also utilized the same five small segments on the cation-binding MIDAS face for ligand engagement. The hypothesis to be tested in Aim I is that different contact residues within these five segments contact these structurally-unrelated ligands. This mosaic model of ligand binding to alphaMbeta2 will be tested by systematic mutagenesis involving not only loss but also gain-in-function strategies. Preliminary data in the application indicate that P2, a 19 amino acid peptide corresponding to residues 377-395 of the gamma-chain of fibrinogen, contains all the information needed to support alphaMbeta2 -dependent cell adhesion, migration and activation.
In Aim 2, the role of the individual amino acids within P2 and within alphaMbeta2 in eliciting these responses will be dissected. The hypothesis to be tested is that, in order to generate the differential intracellular signals required to elicit these alphaMbeta2 -dependent cellular responses, different contact residues in the peptide and in the receptor must interact.
Aim 3 focuses upon the interaction of the major opportunistic pathogen C. albicans with alphaMbeta2. Evidence for release of a soluble ligand, CASL, from the fungus, which interacts with alphaMbeta2 to induce cell adhesion and migration, has been developed. CASL will be isolated, its sequence determined, and its role in controlling the virulence of C. albicans will be established in vitro and in vivo. Overall, the proposed studies have the potential to provide fundamental insights into the functioning of a receptor, alphaMbeta2, which plays a major role in controlling leukocyte responses. The insights gained from these studies are likely to extend to other leukocyte integrins as well. Specific therapeutic targets for controlling the pathogenesis associated with insufficient or overly exuberant alphaMbeta2 -mediated responses, ranging from inflammatory responses to fungal infections, will also be identified through these studies.
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