title: Positive modulation of IL-12 signaling by sphingosine kinase 2 associating with the IL-12 receptor beta 1 cytoplasmic region.
abstract: IL-12 is a key immunoregulatory cytokine that promotes Th1 differentiation and cell-mediated immune responses. IL-12 stimulation results in the activation of Janus kinase 2 and tyrosine kinase 2 and, subsequently, STAT4 and STAT3. In addition, mitogen-activated protein kinase kinase 6/p38 mitogen-activated protein kinase and phosphatidylinositol 3-kinase/Akt pathways have been recently demonstrated to be activated by IL-12 and play an important role in IL-12 signaling. To further elucidate the molecular mechanism underlying IL-12 signaling, we have performed a yeast two-hybrid screening and identified mouse sphingosine kinase 2 (SPHK2) as a molecule associating with the mouse IL-12Rbeta1 cytoplasmic region. Analyses of various mutants of each molecule revealed that the region including the proline-rich domain in SPHK2 is probably responsible for the binding to IL-12Rbeta1, while the regions including the carboxyl terminus and Box II in the IL-12Rbeta1 cytoplasmic region appear to be involved in the binding to SPHK2. Transient expression of wild-type SPHK2 in T cell hybridoma augmented IL-12-induced STAT4-mediated transcriptional activation. Ectopic expression of dominant-negative SPHK2 in Th1 cell clone significantly reduced IL-12-induced IFN-gamma production, while that of wild-type SPHK2 enhanced it. In contrast, the expression minimally affected IL-12-induced proliferation. A similar decrease in IL-12-induced IFN-gamma production was observed when dominant-negative SPHK2 was expressed in activated primary T cells using a retroviral expression system. These results suggest that SPHK2 associates with the IL-12Rbeta1 cytoplasmic region and probably plays a role in modulating IL-12 signaling.
authors.affiliation: Intractable Disease Research Center, Tokyo Medical University, Tokyo, Japan. yoshimot@tokyo-med.ac.jp
authors.forename: Takayuki
authors.initials: T
authors.lastname: Yoshimoto
authors.forename: Masae
authors.initials: M
authors.lastname: Furuhata
authors.forename: Sadahiro
authors.initials: S
authors.lastname: Kamiya
authors.forename: Masayuki
authors.initials: M
authors.lastname: Hisada
authors.forename: Hiroko
authors.initials: H
authors.lastname: Miyaji
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Magami
authors.forename: Koh
authors.initials: K
authors.lastname: Yamamoto
authors.forename: Hiromi
authors.initials: H
authors.lastname: Fujiwara
authors.forename: Junichiro
authors.initials: J
authors.lastname: Mizuguchi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D002999:Clone Cells; D003593:Cytoplasm / Q000201:enzymology* / Q000235:genetics / Q000276:immunology*; D004268:DNA-Binding Proteins / Q000378:metabolism; D015536:Down-Regulation / Q000235:genetics / Q000276:immunology; D020558:GTP Phosphohydrolases / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D015967:Gene Expression Regulation, Viral / Q000276:immunology; D006801:Humans; D006825:Hybridomas; D007371:Interferon-gamma / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D018664:Interleukin-12 / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D008213:Lymphocyte Activation / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D010449:Peptide Mapping; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D017434:Protein Structure, Tertiary / Q000235:genetics; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism*; D018123:Receptors, Interleukin / Q000235:genetics / Q000378:metabolism*; D053707:Receptors, Interleukin-12; D012190:Retroviridae / Q000201:enzymology / Q000276:immunology; D050797:STAT4 Transcription Factor; D015398:Signal Transduction / Q000276:immunology*; D013601:T-Lymphocytes / Q000201:enzymology / Q000276:immunology / Q000821:virology; D018417:Th1 Cells / Q000201:enzymology / Q000276:immunology; D015534:Trans-Activators / Q000378:metabolism; D015533:Transcriptional Activation / Q000276:immunology; D014407:Tumor Cells, Cultured; D020798:Two-Hybrid System Techniques; D015854:Up-Regulation / Q000235:genetics / Q000276:immunology; D000099157:Sphingosine Kinase
chemical_list: D004268:DNA-Binding Proteins; D020558:GTP Phosphohydrolases; D007371:Interferon-gamma; D018664:Interleukin-12; D007527:Isoenzymes; D009687:Nuclear Proteins; D017853:Phosphotransferases (Alcohol Group Acceptor); D021122:Protein Subunits; D018123:Receptors, Interleukin; D053707:Receptors, Interleukin-12; D050797:STAT4 Transcription Factor; D015534:Trans-Activators; D000099157:Sphingosine Kinase; C494090:STAT4 protein, human; C494091:Stat4 protein, mouse; C477973:Gvin1 protein, mouse