Association of intercellular adhesion molecule-1 (ICAM-1) with actin-containing cytoskeleton and alpha-actinin.
We have studied the cytoskeletal association of intercellular adhesion molecule-1 (ICAM-1, CD54), an integral membrane protein that functions as a counterreceptor for leukocyte integrins (CD11/CD18). A linkage between ICAM-1 and cytoskeletal elements was suggested by studies showing a different ICAM-1 staining pattern for COS cells transfected with wild-type ICAM-1 or with an ICAM-1 construct that replaces the cytoplasmic and transmembrane domains of ICAM-1 with a glycophosphatidylinositol (GPI) anchor. Wild-type ICAM-1 appeared to localize most prominently in microvilli whereas GPI-ICAM-1 demonstrated a uniform cell surface distribution. Disruption of microfilaments with cytochalasin B (CCB) changed the localization of wild-type ICAM-1 but had no effect on GPI-ICAM-1. Some B-cell lines demonstrated a prominent accumulation of ICAM-1 into the uropod region whereas other cell surface proteins examined were not preferentially localized. CCB also induced redistribution of ICAM-1 in these cells. For characterization of cytoskeletal proteins interacting with ICAM-1, a 28-residue peptide that encompasses the entire predicted cytoplasmic domain (ICAM-1,478-505) was synthesized, coupled to Sepharose-4B, and used as an affinity matrix. One of the most predominant proteins eluted either with soluble ICAM-1,478-505-peptide or EDTA, was 100 kD, had a pI of 5.5, and in Western blots reacted with alpha-actinin antibodies. A direct association between alpha-actinin and ICAM-1 was demonstrated by binding of purified alpha-actinin to ICAM-1,478-505-peptide and to immunoaffinity purified ICAM-1 and by a strict colocalization of ICAM-1 with alpha-actinin, but not with the cytoskeletal proteins talin, tensin, and vinculin. The region of ICAM-1,478-505 interacting with alpha-actinin was mapped to the area close to the membrane spanning region. This region contains several positively charged residues and appears to mediate a charged interaction with alpha-actinin which is not highly dependent on the order of the residues.
Carpén
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Center for Blood Research, Harvard Medical School, Boston, Massachusetts 02115.
Pallai
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Staunton
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Springer
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D000185:Actinin / Q000378:metabolism*; D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002460:Cell Line; D002461:Cell Line, Transformed; D003571:Cytochalasin B / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006017:Glycolipids / Q000378:metabolism; D017261:Glycosylphosphatidylinositols; D006801:Humans; D018799:Intercellular Adhesion Molecule-1; D008214:Lymphocytes / Q000378:metabolism; D008969:Molecular Sequence Data; D010716:Phosphatidylinositols / Q000378:metabolism; D014407:Tumor Cells, Cultured
D000199:Actins; D015815:Cell Adhesion Molecules; D006017:Glycolipids; D017261:Glycosylphosphatidylinositols; D010716:Phosphatidylinositols; D000185:Actinin; D018799:Intercellular Adhesion Molecule-1; D003571:Cytochalasin B