Ultrastructural examination of cytoskeletal linkage of L-selectin and comparison of L-selectin cytoskeletal association to that of other human and bovine lymphocyte surface antigens.
L-selectin is constitutively expressed on most leukocytes and is responsible for the initial events in cell trafficking termed tethering and rolling. Recently, L-selectin has been shown to associate with the actin-based cytoskeleton under a variety of conditions. In an effort to better understand L-selectin cytoskeletal association and the ultrastructural nature of the cytoskeleton itself, we provide a comparison of the cytoskeletal association of various human and bovine surface proteins in relation to L-selectin. Electron microscopic examination of the cytoskeleton provided further data on the ultrastructure of freshly isolated peripheral lymphocytes as well as demonstrated L-selectin localization to the periphery of the cytoskeleton following low dose detergent treatment of the cell. Clusters of colloidal-gold-stained L-selectin were found on the surface of the detergent-treated lymphocytes, even though these particles completely lacked microvilli. By flow cytometry, we have defined three distinct patterns of cytoskeletal association; constitutive, inductive, and mAb crosslink-induced, and assigned human and bovine CD2, CD3, CD4, CD5, CD8, CD18, CD19, CD44, CD45RA, CD45RO, alphabeta TCR, gammadelta TCR, E-selectin ligands, and L-selectin surface antigens to one of these respective patterns. SDS-PAGE analyses confirmed most of the flow cytometry results. Depending upon its conformation, L-selectin fell into the inductive or mAb crosslink-induced pattern of association, similar to E-selectin ligand(s). Our data provide additional insight into the functional role of L-selectin and the cytoskeleton in immunological events.
Leid
Jeff G
JG
Veterinary Molecular Biology, Montana State University, Bozeman 59717, USA. jleid@erc.montana.edu
Speer
C A
CA
Jutila
Mark A
MA
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism; D000954:Antigens, Surface / Q000276:immunology / Q000378:metabolism*; D002417:Cattle; D003599:Cytoskeleton / Q000378:metabolism* / Q000648:ultrastructure; D003902:Detergents / Q000737:chemistry; D005434:Flow Cytometry; D006801:Humans; D019041:L-Selectin / Q000378:metabolism*; D008214:Lymphocytes / Q000276:immunology* / Q000378:metabolism; D011948:Receptors, Antigen, T-Cell / Q000276:immunology / Q000378:metabolism; D016176:T-Lymphocyte Subsets / Q000276:immunology / Q000378:metabolism / Q000648:ultrastructure
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D003902:Detergents; D011948:Receptors, Antigen, T-Cell; D019041:L-Selectin