A single amino acid alteration in cytoplasmic domain determines IL-2 promoter activation by ligation of CD28 but not inducible costimulator (ICOS)

J Exp Med. 2003 Jan 20;197(2):257-62. doi: 10.1084/jem.20021305.

Abstract

The CD28 family molecules, CD28, and inducible costimulator (ICOS) all provide positive costimulatory signals. However, unlike CD28, ICOS does not costimulate IL-2 secretion. The YMNM motif that exists in the CD28 cytoplasmic domain is a known binding site for phosphatidylinositol 3-kinase (PI3-K) and Grb2. ICOS possesses the YMFM motif in the corresponding region of CD28 that binds PI3-K but not Grb2. We postulated that the reason that ICOS does not have the ability to induce IL-2 production is because it fails to recruit Grb2. To verify this hypothesis, we generated a mutant ICOS gene that contains the CD28 YMNM motif and measured IL-2 promoter activation after ICOS ligation. The results indicated that ICOS became competent to activate the IL-2 promoter by this single alteration. Further analysis demonstrated that Grb2 binding to ICOS was sufficient to activate the NFAT/AP-1 site in the IL-2 promoter and that the cytoplasmic domain of CD28 outside of the YMNM motif is required for activation of the CD28RE/AP-1 and NF-kappaB sites. Together, these observations lead us to believe that the difference of a single amino acid, which affects Grb2 binding ability, may define a functional difference between the CD28- and ICOS-mediated costimulatory signals.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing*
  • Amino Acid Motifs
  • Amino Acid Substitution
  • Animals
  • Antigens, Differentiation, T-Lymphocyte / chemistry
  • Antigens, Differentiation, T-Lymphocyte / genetics
  • Antigens, Differentiation, T-Lymphocyte / metabolism*
  • Binding Sites / genetics
  • CD28 Antigens / chemistry*
  • CD28 Antigens / genetics
  • CD28 Antigens / metabolism*
  • DNA-Binding Proteins / metabolism
  • GRB2 Adaptor Protein
  • Gene Expression Regulation
  • Humans
  • Inducible T-Cell Co-Stimulator Protein
  • Interleukin-2 / genetics*
  • Jurkat Cells
  • Mice
  • Mutagenesis, Site-Directed
  • NF-kappa B / metabolism
  • NFATC Transcription Factors
  • Nuclear Proteins*
  • Phosphatidylinositol 3-Kinases / metabolism
  • Promoter Regions, Genetic*
  • Protein Structure, Tertiary
  • Proteins / metabolism
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Transcription Factor AP-1 / metabolism
  • Transcription Factors / metabolism
  • Transfection

Substances

  • Adaptor Proteins, Signal Transducing
  • Antigens, Differentiation, T-Lymphocyte
  • CD28 Antigens
  • DNA-Binding Proteins
  • GRB2 Adaptor Protein
  • GRB2 protein, human
  • Grb2 protein, mouse
  • ICOS protein, human
  • Icos protein, mouse
  • Inducible T-Cell Co-Stimulator Protein
  • Interleukin-2
  • NF-kappa B
  • NFATC Transcription Factors
  • Nuclear Proteins
  • Proteins
  • Recombinant Fusion Proteins
  • Transcription Factor AP-1
  • Transcription Factors
  • Phosphatidylinositol 3-Kinases