Actin cytoskeletal association of cytohesin-1 is regulated by specific phosphorylation of its carboxyl-terminal polybasic domain.
Cell adhesion mediated by integrin receptors is controlled by intracellular signal transduction cascades. Cytohesin-1 is an integrin-binding protein and guanine nucleotide exchange factor that activates binding of the leukocyte integrin leukocyte function antigen-1 to its ligand, intercellular adhesion molecule 1. Cytohesin-1 bears a carboxyl-terminal pleckstrin homology domain that aids in reversible membrane recruitment and functional regulation of the protein. Although phosphoinositide-dependent membrane attachment of cytohesin-1 is mediated primarily by the pleckstrin homology domain, this function is further strengthened by a short carboxyl-terminal polybasic amino acid sequence. We show here that a serine/threonine motif within the short polybasic stretch of cytohesin-1 is phosphorylated by purified protein kinase C delta in vitro. Furthermore, the respective residues are also found to be phosphorylated after phorbol ester stimulation in vivo. Biochemical and functional analyses show that phosphorylated cytohesin-1 is able to tightly associate with the actin cytoskeleton, and we further demonstrate that phosphorylation of the protein is required for maximal leukocyte function antigen-1-mediated adhesion of Jurkat cells to intercellular adhesion molecule 1. These data suggest that both phosphatidylinositol 3-kinase and protein kinase C-dependent intracellular pathways that stimulate beta(2)-integrin-mediated adhesion of T lymphocytes converge on cytohesin-1 as functional integrator.
Dierks
H
H
Laboratorium für Molekulare Biologie, Genzentrum der Universität München, Feodor-Lynen-Strasse 25, D-81377 München, Germany.
Kolanus
J
J
Kolanus
W
W
D000199:Actins / Q000378:metabolism*; D000818:Animals; D019556:COS Cells; D002273:Carcinogens / Q000494:pharmacology; D002448:Cell Adhesion / Q000187:drug effects; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D019169:Jurkat Cells; D010766:Phosphorylation / Q000187:drug effects; D017434:Protein Structure, Tertiary; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000199:Actins; D002273:Carcinogens; D015815:Cell Adhesion Molecules; D020662:Guanine Nucleotide Exchange Factors; C100723:cytohesin-1; D018799:Intercellular Adhesion Molecule-1; D013755:Tetradecanoylphorbol Acetate