Analysis of CD28 Cytoplasmic Tail Tyrosine Residues as Regulators and Substrates for the Protein Tyrosine Kinases, EMT and LCK

Philip D. King, Ali Sadra, Joyce M.C. Teng, Xiao Rong Liu, Arnold Han, Annamalai Selvakumar, Avery August, Bo Dupont

Research output: Contribution to journalArticle

56 Citations (Scopus)

Abstract

The CD28 cell surface receptor provides an important costimulatory signal for T cells necessary for their response to Ag. Early events in CD28 signaling include recruitment and activation of phosphatidylinositol 3-kinase (PI3-kinase) and activation of the protein tyrosine kinases (PTKs), LCK and EMT. Recruitment and activation of PI3-kinase is known to be dependent upon phosphorylation of tyrosine 173 of the CD28 cytoplasmic tail contained within a YMNM motif. By contrast, little is known of which residues of the CD28 tail, including tyrosines, are required for the activation of PTKs. To address this we studied the ability of truncation mutants and tyrosine to phenylalanine substitution mutants of the CD28 cytoplasmic tail to activate LCK and EMT in Jurkat T leukemia cells. Our results indicate that 1 ) activation of EMT is partially dependent upon tyrosine 173 of the CD28 tail, although it does not require PI3-kinase activation; 2) activation of LCK is independent of CD28 cytoplasmic tail tyrosine residues; and 3) elements sufficient for the activation of both kinases are contained within the first half of the tail. In addition we studied the CD28 tail as a substrate for both PTKs in in vitro kinase assays. We demonstrate that EMT can phosphorylate all four tyrosines of the CD28 tail, in contrast to LCK, which phosphorylates only tyrosine 173. Together with evidence that in vivo, tyrosines other than tyrosine 173 become phosphorylated following CD28 stimulation, this finding suggests that, like LCK, one function of EMT during CD28 signaling is phosphorylation of the receptor.

Original languageEnglish (US)
Pages (from-to)580-590
Number of pages11
JournalJournal of Immunology
Volume158
Issue number2
StatePublished - Jan 15 1997

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Protein-Tyrosine Kinases
Tyrosine
Phosphatidylinositol 3-Kinase
Phosphotransferases
Phosphorylation
T-Cell Leukemia
Cell Surface Receptors
Phenylalanine
T-Lymphocytes

All Science Journal Classification (ASJC) codes

  • Immunology and Allergy
  • Immunology

Cite this

King, P. D., Sadra, A., Teng, J. M. C., Liu, X. R., Han, A., Selvakumar, A., ... Dupont, B. (1997). Analysis of CD28 Cytoplasmic Tail Tyrosine Residues as Regulators and Substrates for the Protein Tyrosine Kinases, EMT and LCK. Journal of Immunology, 158(2), 580-590.
King, Philip D. ; Sadra, Ali ; Teng, Joyce M.C. ; Liu, Xiao Rong ; Han, Arnold ; Selvakumar, Annamalai ; August, Avery ; Dupont, Bo. / Analysis of CD28 Cytoplasmic Tail Tyrosine Residues as Regulators and Substrates for the Protein Tyrosine Kinases, EMT and LCK. In: Journal of Immunology. 1997 ; Vol. 158, No. 2. pp. 580-590.
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abstract = "The CD28 cell surface receptor provides an important costimulatory signal for T cells necessary for their response to Ag. Early events in CD28 signaling include recruitment and activation of phosphatidylinositol 3-kinase (PI3-kinase) and activation of the protein tyrosine kinases (PTKs), LCK and EMT. Recruitment and activation of PI3-kinase is known to be dependent upon phosphorylation of tyrosine 173 of the CD28 cytoplasmic tail contained within a YMNM motif. By contrast, little is known of which residues of the CD28 tail, including tyrosines, are required for the activation of PTKs. To address this we studied the ability of truncation mutants and tyrosine to phenylalanine substitution mutants of the CD28 cytoplasmic tail to activate LCK and EMT in Jurkat T leukemia cells. Our results indicate that 1 ) activation of EMT is partially dependent upon tyrosine 173 of the CD28 tail, although it does not require PI3-kinase activation; 2) activation of LCK is independent of CD28 cytoplasmic tail tyrosine residues; and 3) elements sufficient for the activation of both kinases are contained within the first half of the tail. In addition we studied the CD28 tail as a substrate for both PTKs in in vitro kinase assays. We demonstrate that EMT can phosphorylate all four tyrosines of the CD28 tail, in contrast to LCK, which phosphorylates only tyrosine 173. Together with evidence that in vivo, tyrosines other than tyrosine 173 become phosphorylated following CD28 stimulation, this finding suggests that, like LCK, one function of EMT during CD28 signaling is phosphorylation of the receptor.",
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King, PD, Sadra, A, Teng, JMC, Liu, XR, Han, A, Selvakumar, A, August, A & Dupont, B 1997, 'Analysis of CD28 Cytoplasmic Tail Tyrosine Residues as Regulators and Substrates for the Protein Tyrosine Kinases, EMT and LCK', Journal of Immunology, vol. 158, no. 2, pp. 580-590.

Analysis of CD28 Cytoplasmic Tail Tyrosine Residues as Regulators and Substrates for the Protein Tyrosine Kinases, EMT and LCK. / King, Philip D.; Sadra, Ali; Teng, Joyce M.C.; Liu, Xiao Rong; Han, Arnold; Selvakumar, Annamalai; August, Avery; Dupont, Bo.

In: Journal of Immunology, Vol. 158, No. 2, 15.01.1997, p. 580-590.

Research output: Contribution to journalArticle

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T1 - Analysis of CD28 Cytoplasmic Tail Tyrosine Residues as Regulators and Substrates for the Protein Tyrosine Kinases, EMT and LCK

AU - King, Philip D.

AU - Sadra, Ali

AU - Teng, Joyce M.C.

AU - Liu, Xiao Rong

AU - Han, Arnold

AU - Selvakumar, Annamalai

AU - August, Avery

AU - Dupont, Bo

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N2 - The CD28 cell surface receptor provides an important costimulatory signal for T cells necessary for their response to Ag. Early events in CD28 signaling include recruitment and activation of phosphatidylinositol 3-kinase (PI3-kinase) and activation of the protein tyrosine kinases (PTKs), LCK and EMT. Recruitment and activation of PI3-kinase is known to be dependent upon phosphorylation of tyrosine 173 of the CD28 cytoplasmic tail contained within a YMNM motif. By contrast, little is known of which residues of the CD28 tail, including tyrosines, are required for the activation of PTKs. To address this we studied the ability of truncation mutants and tyrosine to phenylalanine substitution mutants of the CD28 cytoplasmic tail to activate LCK and EMT in Jurkat T leukemia cells. Our results indicate that 1 ) activation of EMT is partially dependent upon tyrosine 173 of the CD28 tail, although it does not require PI3-kinase activation; 2) activation of LCK is independent of CD28 cytoplasmic tail tyrosine residues; and 3) elements sufficient for the activation of both kinases are contained within the first half of the tail. In addition we studied the CD28 tail as a substrate for both PTKs in in vitro kinase assays. We demonstrate that EMT can phosphorylate all four tyrosines of the CD28 tail, in contrast to LCK, which phosphorylates only tyrosine 173. Together with evidence that in vivo, tyrosines other than tyrosine 173 become phosphorylated following CD28 stimulation, this finding suggests that, like LCK, one function of EMT during CD28 signaling is phosphorylation of the receptor.

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