Dynamic association of L-selectin with the lymphocyte cytoskeletal matrix.
L-selectin mediates lymphocyte extravasation into lymphoid tissues through binding to sialomucin-like receptors on the surface of high endothelial venules (HEV). This study examines the biochemical basis and regulation of interactions between L-selectin, an integral transmembrane protein, and the lymphocyte cytoskeleton. Using a detergent-based extraction procedure, constitutive associations between L-selectin and the insoluble cytoskeletal matrix could not be detected. However, engagement of the L-selectin lectin domain by Abs or by glycosylation-dependent cell adhesion molecule-1, an HEV-derived ligand for L-selectin, rapidly triggered redistribution of L-selectin to the detergent-insoluble cytoskeleton. L-selectin attachment to the cytoskeleton was not prevented by inhibitors of actin/microtubule polymerization (cytochalasin B, colchicine, or nocodozole) or serine/threonine and tyrosine kinase activity (staurosporine, calphostin C, or genistein), although L-selectin-mediated adhesion of human PBL was markedly suppressed by these agents. Exposure of human PBL or murine pre-B transfectants expressing full-length human L-selectin to fever-range hyperthermia also markedly increased L-selectin association with the cytoskeleton, directly correlating with enhanced L-selectin-mediated adhesion. In contrast, a deletion mutant of L-selectin lacking the COOH-terminal 11 amino acids failed to associate with the cytoskeletal matrix in response to Ab cross-linking or hyperthermia stimulation and did not support adhesion to HEV. These studies, when taken together with the previously demonstrated interaction between the L-selectin cytoplasmic domain and the cytoskeletal linker protein alpha-actinin, strongly implicate the actin-based cytoskeleton in dynamically controlling L-selectin adhesion.
Evans
S S
SS
Department of Immunology, Roswell Park Cancer Institute, Buffalo, NY 14263, USA. sevans@sc3101.med.buffalo.edu
Schleider
D M
DM
Bowman
L A
LA
Francis
M L
ML
Kansas
G S
GS
Black
J D
JD
D000911:Antibodies, Monoclonal / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000378:metabolism; D003593:Cytoplasm / Q000502:physiology; D003599:Cytoskeleton / Q000201:enzymology / Q000378:metabolism* / Q000502:physiology; D003902:Detergents; D005334:Fever / Q000276:immunology / Q000378:metabolism; D006801:Humans; D006979:Hyperthermia, Induced; D019041:L-Selectin / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D008214:Lymphocytes / Q000201:enzymology / Q000378:metabolism*; D009077:Mucins / Q000378:metabolism; D011485:Protein Binding / Q000276:immunology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012995:Solubility
D000911:Antibodies, Monoclonal; D003432:Cross-Linking Reagents; D003902:Detergents; D009077:Mucins; D019041:L-Selectin; C075185:sulfated glycoprotein p50; D017346:Protein Serine-Threonine Kinases