The functional interaction of the beta 2 integrin lymphocyte function-associated antigen-1 with junctional adhesion molecule-A is mediated by the I domain.
Binding of the beta(2) integrin LFA-1 (alpha(L)beta(2)) to junctional adhesion molecule-A (JAM-A) has been shown to enhance leukocyte adhesion and transendothelial migration. This is mediated by the membrane-proximal Ig-like domain 2 of JAM-A; however, the location of the JAM-A binding site in LFA-1 has not been identified. We have deleted the I domain in the alpha(L) subunit of LFA-1 and expressed this alpha(L) mutant in alpha(l)-deficient Jurkat J-beta(2).7 cells to demonstrate that the I domain of LFA-1 is crucial for their adhesion to immobilized JAM-A. This was substantiated by blocking the stimulated adhesion of wild-type Jurkat T cells or monocytic Mono Mac 6 cells to JAM-A using the I domain-directed mAb TS1/22 or the small molecule antagonist BIRT 377, which stabilizes the low-affinity conformation of the I domain. The immobilized LFA-1 I domain locked in the open high-affinity conformation was sufficient to support binding of transfected Chinese hamster ovary cells expressing JAM-A. Solid-phase binding assays confirmed a direct interaction of recombinant JAM-A with the immobilized locked-open I domain. These data provide the first evidence that the I domain of LFA-1 contains a functional binding site for JAM-A.
Fraemohs
Line
L
Department of Molecular Cardiovascular Research, Rheinisch-Westfälische Technische Hochschule, University Hospital Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany.
Koenen
Rory R
RR
Ostermann
Georg
G
Heinemann
Bo
B
Weber
Christian
C
D000818:Animals; D019138:Antibodies, Blocking / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001667:Binding, Competitive / Q000235:genetics / Q000276:immunology; D018821:CD18 Antigens / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D016466:CHO Cells; D015815:Cell Adhesion Molecules / Q000378:metabolism* / Q000502:physiology; D006224:Cricetinae; D006801:Humans; D048289:Imidazolidines / Q000494:pharmacology; D062726:Junctional Adhesion Molecules; D019169:Jurkat Cells; D016169:Lymphocyte Function-Associated Antigen-1 / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics / Q000502:physiology; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D017384:Sequence Deletion
D019138:Antibodies, Blocking; D000911:Antibodies, Monoclonal; C400461:BIRT 377; D018821:CD18 Antigens; D015815:Cell Adhesion Molecules; D048289:Imidazolidines; D062726:Junctional Adhesion Molecules; D016169:Lymphocyte Function-Associated Antigen-1; D011994:Recombinant Proteins