title: Functional mapping of the cytoplasmic region of intercellular adhesion molecule-3 reveals important roles for serine residues.
abstract: Intercellular adhesion molecule-3 (ICAM-3), a ligand for beta2 integrins, elicits a variety of activation responses in lymphocytes. We describe a functional mapping study that focuses on the 37-residue cytoplasmic region of ICAM-3. Carboxyl-terminal truncations delineated portions involved in T cell antigen receptor costimulation, homotypic aggregation, and cellular spreading. Truncation of the membrane distal 25 residues resulted in loss of T cell antigen receptor costimulation as determined by interleukin 2 secretion. Aggregation and cell spreading were sensitive to truncation of the membrane distal and proximal thirds of the cytoplasmic portion. Phosphoamino acid analysis revealed that ICAM-3 from activated cells contained phosphoserine and phosphopeptide mapping identified Ser489 as a site of phosphorylation in vivo. Mutation of Ser489 or Ser515 to alanine blocked interleukin 2 secretion, aggregation and cell spreading, while mutation of other serine residues affected only a subset of functions. Ser489 was a phosphorylation site in vitro for recombinant protein kinase Ctheta. Finally, treatment of Jurkat cells with chelerythrine chloride, a protein kinase C inhibitor, prevented ICAM-3-triggered spreading. This study delineates separable regions and amino acid residues within the cytoplasmic portion of ICAM-3 that are important for T cell function.
authors.affiliation: ICOS Corporation, Bothell, Washington 98021, USA. jhayflick@icos.com
authors.forename: J S
authors.initials: JS
authors.lastname: Hayflick
authors.forename: J
authors.initials: J
authors.lastname: Stine
authors.forename: R
authors.initials: R
authors.lastname: Fox
authors.forename: D
authors.initials: D
authors.lastname: Hoekstra
authors.forename: W M
authors.initials: WM
authors.lastname: Gallatin
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD; D000943:Antigens, Differentiation; D001665:Binding Sites; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules / Q000737:chemistry* / Q000378:metabolism; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D018014:Gene Transfer Techniques; D006801:Humans; D019169:Jurkat Cells; D051379:Mice; D008969:Molecular Sequence Data; D012694:Serine / Q000737:chemistry* / Q000378:metabolism
chemical_list: D015703:Antigens, CD; D000943:Antigens, Differentiation; D015815:Cell Adhesion Molecules; C483616:ICAM3 protein, human; D012694:Serine