Molecular Basis of MO1/LFA-1 Deficiency

Massachusetts General Hospital, Boston MA

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Abstract

(32 integrins are a|3 heterodimeric receptors that mediate bidirectional signaling in leukocytes, which underlies most functions,mediated by these cells including leukocyte adhesion, migration, proliferation and immune clearance. (32 integrins are kept in a default low affinity state on the leukocyte surface, and normally switch to the high affinity ligand-binding state in response to activating inflammatory mediators. Dysregulation of these receptors is associated with many inflammatory pathologies due to uncontrolled cell adhesion. Thus understanding how these adhesion receptors are regulated is essential in promoting health and preventing organ injury. A primary goal of this grant since its inception has been to derive structure-activity relationships in these receptors to better understand their role in leukocyte biology and to translate some of the advances in structure into potential therapies. Advances achieved during the current funding cycleincluded the derivation of the structure of the ligand binding domain (aA) of the (32 integrin CDllb/CD18 in complex with a ligand- mimetic Fab, structure determination of the complete genuflexed ectodomain of an ccA-lacking integrin, and the identification of three allosteric sites in (32 integrins that regulate its activation state. A challenge now is to relate some of these findings to the receptor in its native membrane-bound state and to determine if the insights obtained from the structural data can be translated into useful therapies. Our preliminary studies now show that the quaternary relationships of the ligand binding aA domain of membrane-bound CDllb/CD18 can be visualized using fluorescencelifetime imaging microscopy, that novel reagents can be used to probe the receptor in live cells in its active and inactive states and that the active state of aA is renoprotective in a model of renal inflammation. Consequently, four aims are proposed that build on these observations using biochemical, biophysical, crystallographic and genetic approaches in addition to murine models of renal inflammation.