Abstract
Immunoreceptor tyrosine-based inhibitory motifs (ITIM) have been implicated in the negative modulation of immunoreceptor signaling pathways. The IL-4R alpha-chain (IL-4Ralpha) contains a putative ITIM in the carboxyl terminal. To determine the role of ITIM in the IL-4 signaling pathway, we ablated the ITIM of IL-4Ralpha by deletion and site-directed mutagenesis and stably expressed the wild-type (WT) and mutant hIL-4Ralpha in 32D/insulin receptor substrate-2 (IRS-2) cells. Strikingly, 32D/IRS-2 cells expressing mutant human (h)IL-4Ralpha were hyperproliferative in response to IL-4 compared with cells expressing WT hIL-4Ralpha. Enhanced tyrosine phosphorylation of Stat6, but not IRS-2, induced by hIL-4 was observed in cells expressing mutant Y713F. Using peptides corresponding to the ITIM of hIL-4Ralpha, we demonstrate that tyrosine-phosphorylated peptides, but not their nonphosphorylated counterparts, coprecipitate SH2-containing tyrosine phosphatase-1, SH2-containing tyrosine phosphatase-2, and SH2-containing inositol 5'-phosphatase. The in vivo association of SH2-containing inositol 5'-phosphatase with IL-4Ralpha was verified by coimmunoprecipitation with anti-IL-4Ralpha Abs. These results demonstrate a functional role for ITIM in the regulation of IL-4-induced proliferation.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Amino Acid Motifs / genetics
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Amino Acid Sequence
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Cell Line
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Cytoplasm / enzymology
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Cytoplasm / genetics
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Cytoplasm / immunology
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Enzyme Activation / genetics
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Enzyme Activation / immunology
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Humans
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Insulin Receptor Substrate Proteins
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Interleukin-4 / physiology*
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Intracellular Signaling Peptides and Proteins
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Janus Kinase 1
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Lymphocyte Activation / genetics
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Lymphocyte Activation / immunology*
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Molecular Sequence Data
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Mutagenesis, Site-Directed
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Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases
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Phosphoproteins / metabolism
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Phosphoric Monoester Hydrolases / metabolism*
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Phosphoric Monoester Hydrolases / physiology
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Phosphorylation
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Protein Phosphatase 1
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Protein Phosphatase 2
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Protein Tyrosine Phosphatase, Non-Receptor Type 11
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Protein Tyrosine Phosphatase, Non-Receptor Type 6
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Protein Tyrosine Phosphatases / metabolism
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Protein-Tyrosine Kinases / metabolism
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Receptors, Immunologic / genetics
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Receptors, Immunologic / metabolism*
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Receptors, Immunologic / physiology
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Receptors, Interleukin-4 / genetics
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Receptors, Interleukin-4 / metabolism*
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Receptors, Interleukin-4 / physiology
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SH2 Domain-Containing Protein Tyrosine Phosphatases
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STAT6 Transcription Factor
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Sequence Deletion
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Signal Transduction / genetics
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Signal Transduction / immunology
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Trans-Activators / metabolism
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Tyrosine / genetics
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Tyrosine / metabolism*
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src Homology Domains / immunology*
Substances
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IRS2 protein, human
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Insulin Receptor Substrate Proteins
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Intracellular Signaling Peptides and Proteins
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Phosphoproteins
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Receptors, Immunologic
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Receptors, Interleukin-4
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STAT6 Transcription Factor
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STAT6 protein, human
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Trans-Activators
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Interleukin-4
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Tyrosine
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Protein-Tyrosine Kinases
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JAK1 protein, human
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Janus Kinase 1
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Protein Phosphatase 1
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Protein Phosphatase 2
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Phosphoric Monoester Hydrolases
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PTPN11 protein, human
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PTPN6 protein, human
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Protein Tyrosine Phosphatase, Non-Receptor Type 11
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Protein Tyrosine Phosphatase, Non-Receptor Type 6
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Protein Tyrosine Phosphatases
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SH2 Domain-Containing Protein Tyrosine Phosphatases
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INPPL1 protein, human
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Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases