Abstract
TRAC-1 (T cell RING (really interesting new gene) protein identified in activation screen) is a novel E3 ubiquitin ligase identified from a retroviral vector-based T cell surface activation marker screen. The C-terminal truncated TRAC-1 specifically inhibited anti-TCR-mediated CD69 up-regulation in Jurkat cells, a human T leukemic cell line. In this study, we show that TRAC-1 is a RING finger ubiquitin E3 ligase with highest expression in lymphoid tissues. Point mutations that disrupt the Zn(2+)-chelating ability of its amino-terminal RING finger domain abolished TRAC-1's ligase activity and the dominant inhibitory effect of C-terminal truncated TRAC-1 on TCR stimulation. The results of in vitro biochemical studies indicate that TRAC-1 can stimulate the formation of both K48- and K63-linked polyubiquitin chains and therefore could potentially activate both degradative and regulatory ubiquitin-dependent pathways. Antisense oligonucleotides to TRAC-1 specifically reduced TRAC-1 mRNA levels in Jurkat and primary T cells and inhibited their activation in response to TCR cross-linking. Collectively, these results indicate that the E3 ubiquitin ligase TRAC-1 functions as a positive regulator of T cell activation.
MeSH terms
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Amino Acid Motifs
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Amino Acid Sequence
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Base Sequence
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Catalysis
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Cell Cycle Proteins / biosynthesis
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / isolation & purification
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Cell Cycle Proteins / physiology*
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Cell Line
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Cell Line, Tumor
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DNA-Binding Proteins
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Humans
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Jurkat Cells
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Lymphocyte Activation / genetics
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Lymphocyte Activation / immunology*
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Lymphoid Tissue / cytology
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Lymphoid Tissue / enzymology
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Lymphoid Tissue / immunology
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Molecular Sequence Data
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Nuclear Proteins / biosynthesis
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Nuclear Proteins / genetics
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Nuclear Proteins / isolation & purification
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Nuclear Proteins / physiology*
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Nuclear Receptor Co-Repressor 2
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Peptide Fragments / genetics
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Peptide Fragments / metabolism
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Polyubiquitin / metabolism
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Receptors, Antigen, T-Cell / physiology
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Repressor Proteins
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Saccharomyces cerevisiae Proteins / metabolism
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Signal Transduction / genetics
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Signal Transduction / immunology
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T-Lymphocytes / enzymology*
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T-Lymphocytes / immunology*
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T-Lymphocytes / metabolism
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Transcription Factors / metabolism
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Ubiquitin-Conjugating Enzymes / metabolism
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Ubiquitin-Protein Ligases / biosynthesis
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Ubiquitin-Protein Ligases / genetics
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Ubiquitin-Protein Ligases / isolation & purification
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Ubiquitin-Protein Ligases / physiology*
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Up-Regulation / genetics
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Up-Regulation / immunology*
Substances
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Cell Cycle Proteins
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DNA-Binding Proteins
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NCOR2 protein, human
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Nuclear Proteins
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Nuclear Receptor Co-Repressor 2
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Peptide Fragments
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Receptors, Antigen, T-Cell
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Repressor Proteins
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Saccharomyces cerevisiae Proteins
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Transcription Factors
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Polyubiquitin
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UBC13 protein, S cerevisiae
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UBE2D3 protein, human
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UBE2V1 protein, human
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Ubiquitin-Conjugating Enzymes
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Ubiquitin-Protein Ligases