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. 2002 Apr 1;363(Pt 1):175–182. doi: 10.1042/0264-6021:3630175

Opposing roles of serine/threonine kinases MEKK1 and LOK in regulating the CD28 responsive element in T-cells.

Li Tao 1, Scott Wadsworth 1, Jason Mercer 1, Cynthia Mueller 1, Kirsten Lynn 1, John Siekierka 1, Avery August 1
PMCID: PMC1222464  PMID: 11903060

Abstract

T-cell activation requires signals from both the T-cell receptor (TcR) and other co-stimulatory molecules such as CD28. TcR- and CD28-mediated signals are integrated during T-cell activation resulting in the expression of cytokine genes such as interleukin-2 (IL-2). An enhancer element (CD28RE) of the IL-2 gene specifically responsive to CD28 signals has been previously identified and characterized. This response element and an adjacent Activated Protein-1 (nuclear factor-interleukin-2B) site together (RE/AP1) were shown to complex with c-rel, AP-1 and other factors. However, details of the signal transduction pathways leading from CD28 to the composite response element remain poorly understood. We present data showing that overexpression of the serine threonine kinase, mitogen-activated protein kinase/extracellular-signal-regulated kinase kinase kinase-1 (MEKK1), but not nuclear factor-kappa B inducing kinase, or MAP kinase/ERK kinase-1 (MEK1), can significantly increase the level of CD28RE/AP1-driven luciferase (Luc) reporter gene expression in Jurkat E6-1 cells. A MEKK1 dominant negative mutant blocked such activation induced by stimulation with Raji B cells and the superantigen staphylococcus enterotoxin E (SEE), as well as via CD3/CD28. Mutations in either site of the RE/AP1 element abolished MEKK1-induced Luc expression. Calcineurin inhibitors, CsA and FK520, or inhibitors of p38 kinase (SB 203580), or MEK1 (PD 098059), did not affect MEKK1-induced reporter activation. These results directly implicate MEKK1 in the CD28 signalling pathway that activates the CD28 response element. Co-expression of the lymphocyte-oriented kinase (LOK) kinase attenuated Raji/SEE-induced IL-2 production in Jurkat cells, as well as MEKK1 and Raji/SEE-induced reporter gene activation. These data suggest that MEKK1 and LOK may have opposing roles in regulating the CD28RE/AP1 element.

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Selected References

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  1. August A., Dupont B. CD28 of T lymphocytes associates with phosphatidylinositol 3-kinase. Int Immunol. 1994 May;6(5):769–774. doi: 10.1093/intimm/6.5.769. [DOI] [PubMed] [Google Scholar]
  2. August A., Gibson S., Kawakami Y., Kawakami T., Mills G. B., Dupont B. CD28 is associated with and induces the immediate tyrosine phosphorylation and activation of the Tec family kinase ITK/EMT in the human Jurkat leukemic T-cell line. Proc Natl Acad Sci U S A. 1994 Sep 27;91(20):9347–9351. doi: 10.1073/pnas.91.20.9347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Barz C., Nagel T., Truitt K. E., Imboden J. B. Mutational analysis of CD28-mediated costimulation of Jun-N-terminal kinase and IL-2 production. J Immunol. 1998 Nov 15;161(10):5366–5372. [PubMed] [Google Scholar]
  4. Boguski M. S., Lowe T. M., Tolstoshev C. M. dbEST--database for "expressed sequence tags". Nat Genet. 1993 Aug;4(4):332–333. doi: 10.1038/ng0893-332. [DOI] [PubMed] [Google Scholar]
  5. Butscher W. G., Powers C., Olive M., Vinson C., Gardner K. Coordinate transactivation of the interleukin-2 CD28 response element by c-Rel and ATF-1/CREB2. J Biol Chem. 1998 Jan 2;273(1):552–560. doi: 10.1074/jbc.273.1.552. [DOI] [PubMed] [Google Scholar]
  6. Chambers C. A., Allison J. P. Co-stimulation in T cell responses. Curr Opin Immunol. 1997 Jun;9(3):396–404. doi: 10.1016/s0952-7915(97)80087-8. [DOI] [PubMed] [Google Scholar]
  7. Diener K., Wang X. S., Chen C., Meyer C. F., Keesler G., Zukowski M., Tan T. H., Yao Z. Activation of the c-Jun N-terminal kinase pathway by a novel protein kinase related to human germinal center kinase. Proc Natl Acad Sci U S A. 1997 Sep 2;94(18):9687–9692. doi: 10.1073/pnas.94.18.9687. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Faris M., Kokot N., Lee L., Nel A. E. Regulation of interleukin-2 transcription by inducible stable expression of dominant negative and dominant active mitogen-activated protein kinase kinase kinase in jurkat T cells. Evidence for the importance of Ras in a pathway that is controlled by dual receptor stimulation. J Biol Chem. 1996 Nov 1;271(44):27366–27373. doi: 10.1074/jbc.271.44.27366. [DOI] [PubMed] [Google Scholar]
  9. Fraser J. D., Irving B. A., Crabtree G. R., Weiss A. Regulation of interleukin-2 gene enhancer activity by the T cell accessory molecule CD28. Science. 1991 Jan 18;251(4991):313–316. doi: 10.1126/science.1846244. [DOI] [PubMed] [Google Scholar]
  10. Fraser J. D., Newton M. E., Weiss A. CD28 and T cell antigen receptor signal transduction coordinately regulate interleukin 2 gene expression in response to superantigen stimulation. J Exp Med. 1992 Apr 1;175(4):1131–1134. doi: 10.1084/jem.175.4.1131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fraser J. D., Straus D., Weiss A. Signal transduction events leading to T-cell lymphokine gene expression. Immunol Today. 1993 Jul;14(7):357–362. doi: 10.1016/0167-5699(93)90236-E. [DOI] [PubMed] [Google Scholar]
  12. Ghosh P., Tan T. H., Rice N. R., Sica A., Young H. A. The interleukin 2 CD28-responsive complex contains at least three members of the NF kappa B family: c-Rel, p50, and p65. Proc Natl Acad Sci U S A. 1993 Mar 1;90(5):1696–1700. doi: 10.1073/pnas.90.5.1696. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Harding F. A., McArthur J. G., Gross J. A., Raulet D. H., Allison J. P. CD28-mediated signalling co-stimulates murine T cells and prevents induction of anergy in T-cell clones. Nature. 1992 Apr 16;356(6370):607–609. doi: 10.1038/356607a0. [DOI] [PubMed] [Google Scholar]
  14. Harhaj E. W., Sun S. C. IkappaB kinases serve as a target of CD28 signaling. J Biol Chem. 1998 Sep 25;273(39):25185–25190. doi: 10.1074/jbc.273.39.25185. [DOI] [PubMed] [Google Scholar]
  15. Hu M. C., Qiu W. R., Wang X., Meyer C. F., Tan T. H. Human HPK1, a novel human hematopoietic progenitor kinase that activates the JNK/SAPK kinase cascade. Genes Dev. 1996 Sep 15;10(18):2251–2264. doi: 10.1101/gad.10.18.2251. [DOI] [PubMed] [Google Scholar]
  16. Jain J., Loh C., Rao A. Transcriptional regulation of the IL-2 gene. Curr Opin Immunol. 1995 Jun;7(3):333–342. doi: 10.1016/0952-7915(95)80107-3. [DOI] [PubMed] [Google Scholar]
  17. June C. H., Bluestone J. A., Nadler L. M., Thompson C. B. The B7 and CD28 receptor families. Immunol Today. 1994 Jul;15(7):321–331. doi: 10.1016/0167-5699(94)90080-9. [DOI] [PubMed] [Google Scholar]
  18. Kaga S., Ragg S., Rogers K. A., Ochi A. Activation of p21-CDC42/Rac-activated kinases by CD28 signaling: p21-activated kinase (PAK) and MEK kinase 1 (MEKK1) may mediate the interplay between CD3 and CD28 signals. J Immunol. 1998 May 1;160(9):4182–4189. [PubMed] [Google Scholar]
  19. Kaga S., Ragg S., Rogers K. A., Ochi A. Stimulation of CD28 with B7-2 promotes focal adhesion-like cell contacts where Rho family small G proteins accumulate in T cells. J Immunol. 1998 Jan 1;160(1):24–27. [PubMed] [Google Scholar]
  20. Karin M., Liu Z. g., Zandi E. AP-1 function and regulation. Curr Opin Cell Biol. 1997 Apr;9(2):240–246. doi: 10.1016/s0955-0674(97)80068-3. [DOI] [PubMed] [Google Scholar]
  21. Kempiak S. J., Hiura T. S., Nel A. E. The Jun kinase cascade is responsible for activating the CD28 response element of the IL-2 promoter: proof of cross-talk with the I kappa B kinase cascade. J Immunol. 1999 Mar 15;162(6):3176–3187. [PubMed] [Google Scholar]
  22. Kuramochi S., Moriguchi T., Kuida K., Endo J., Semba K., Nishida E., Karasuyama H. LOK is a novel mouse STE20-like protein kinase that is expressed predominantly in lymphocytes. J Biol Chem. 1997 Sep 5;272(36):22679–22684. doi: 10.1074/jbc.272.36.22679. [DOI] [PubMed] [Google Scholar]
  23. Köntgen F., Grumont R. J., Strasser A., Metcalf D., Li R., Tarlinton D., Gerondakis S. Mice lacking the c-rel proto-oncogene exhibit defects in lymphocyte proliferation, humoral immunity, and interleukin-2 expression. Genes Dev. 1995 Aug 15;9(16):1965–1977. doi: 10.1101/gad.9.16.1965. [DOI] [PubMed] [Google Scholar]
  24. Lai J. H., Horvath G., Subleski J., Bruder J., Ghosh P., Tan T. H. RelA is a potent transcriptional activator of the CD28 response element within the interleukin 2 promoter. Mol Cell Biol. 1995 Aug;15(8):4260–4271. doi: 10.1128/mcb.15.8.4260. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Leberer E., Dignard D., Harcus D., Thomas D. Y., Whiteway M. The protein kinase homologue Ste20p is required to link the yeast pheromone response G-protein beta gamma subunits to downstream signalling components. EMBO J. 1992 Dec;11(13):4815–4824. doi: 10.1002/j.1460-2075.1992.tb05587.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Lee F. S., Hagler J., Chen Z. J., Maniatis T. Activation of the IkappaB alpha kinase complex by MEKK1, a kinase of the JNK pathway. Cell. 1997 Jan 24;88(2):213–222. doi: 10.1016/s0092-8674(00)81842-5. [DOI] [PubMed] [Google Scholar]
  27. Lee F. S., Peters R. T., Dang L. C., Maniatis T. MEKK1 activates both IkappaB kinase alpha and IkappaB kinase beta. Proc Natl Acad Sci U S A. 1998 Aug 4;95(16):9319–9324. doi: 10.1073/pnas.95.16.9319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lenschow D. J., Walunas T. L., Bluestone J. A. CD28/B7 system of T cell costimulation. Annu Rev Immunol. 1996;14:233–258. doi: 10.1146/annurev.immunol.14.1.233. [DOI] [PubMed] [Google Scholar]
  29. Lin X., Cunningham E. T., Jr, Mu Y., Geleziunas R., Greene W. C. The proto-oncogene Cot kinase participates in CD3/CD28 induction of NF-kappaB acting through the NF-kappaB-inducing kinase and IkappaB kinases. Immunity. 1999 Feb;10(2):271–280. doi: 10.1016/s1074-7613(00)80027-8. [DOI] [PubMed] [Google Scholar]
  30. Malinin N. L., Boldin M. P., Kovalenko A. V., Wallach D. MAP3K-related kinase involved in NF-kappaB induction by TNF, CD95 and IL-1. Nature. 1997 Feb 6;385(6616):540–544. doi: 10.1038/385540a0. [DOI] [PubMed] [Google Scholar]
  31. McGuire K. L., Iacobelli M. Involvement of Rel, Fos, and Jun proteins in binding activity to the IL-2 promoter CD28 response element/AP-1 sequence in human T cells. J Immunol. 1997 Aug 1;159(3):1319–1327. [PubMed] [Google Scholar]
  32. Nishina H., Bachmann M., Oliveira-dos-Santos A. J., Kozieradzki I., Fischer K. D., Odermatt B., Wakeham A., Shahinian A., Takimoto H., Bernstein A. Impaired CD28-mediated interleukin 2 production and proliferation in stress kinase SAPK/ERK1 kinase (SEK1)/mitogen-activated protein kinase kinase 4 (MKK4)-deficient T lymphocytes. J Exp Med. 1997 Sep 15;186(6):941–953. doi: 10.1084/jem.186.6.941. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Prasad K. V., Cai Y. C., Raab M., Duckworth B., Cantley L., Shoelson S. E., Rudd C. E. T-cell antigen CD28 interacts with the lipid kinase phosphatidylinositol 3-kinase by a cytoplasmic Tyr(P)-Met-Xaa-Met motif. Proc Natl Acad Sci U S A. 1994 Mar 29;91(7):2834–2838. doi: 10.1073/pnas.91.7.2834. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Ramer S. W., Davis R. W. A dominant truncation allele identifies a gene, STE20, that encodes a putative protein kinase necessary for mating in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1993 Jan 15;90(2):452–456. doi: 10.1073/pnas.90.2.452. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Sabapathy K., Hu Y., Kallunki T., Schreiber M., David J. P., Jochum W., Wagner E. F., Karin M. JNK2 is required for efficient T-cell activation and apoptosis but not for normal lymphocyte development. Curr Biol. 1999 Feb 11;9(3):116–125. doi: 10.1016/s0960-9822(99)80065-7. [DOI] [PubMed] [Google Scholar]
  36. Schafer P. H., Wadsworth S. A., Wang L., Siekierka J. J. p38 alpha mitogen-activated protein kinase is activated by CD28-mediated signaling and is required for IL-4 production by human CD4+CD45RO+ T cells and Th2 effector cells. J Immunol. 1999 Jun 15;162(12):7110–7119. [PubMed] [Google Scholar]
  37. Schlesinger T. K., Fanger G. R., Yujiri T., Johnson G. L. The TAO of MEKK. Front Biosci. 1998 Nov 15;3:D1181–D1186. doi: 10.2741/a354. [DOI] [PubMed] [Google Scholar]
  38. Schneider H., Cai Y. C., Prasad K. V., Shoelson S. E., Rudd C. E. T cell antigen CD28 binds to the GRB-2/SOS complex, regulators of p21ras. Eur J Immunol. 1995 Apr;25(4):1044–1050. doi: 10.1002/eji.1830250428. [DOI] [PubMed] [Google Scholar]
  39. Schwartz R. H. Costimulation of T lymphocytes: the role of CD28, CTLA-4, and B7/BB1 in interleukin-2 production and immunotherapy. Cell. 1992 Dec 24;71(7):1065–1068. doi: 10.1016/s0092-8674(05)80055-8. [DOI] [PubMed] [Google Scholar]
  40. Scott M. L., Fujita T., Liou H. C., Nolan G. P., Baltimore D. The p65 subunit of NF-kappa B regulates I kappa B by two distinct mechanisms. Genes Dev. 1993 Jul;7(7A):1266–1276. doi: 10.1101/gad.7.7a.1266. [DOI] [PubMed] [Google Scholar]
  41. Shapiro V. S., Mollenauer M. N., Weiss A. Nuclear factor of activated T cells and AP-1 are insufficient for IL-2 promoter activation: requirement for CD28 up-regulation of RE/AP. J Immunol. 1998 Dec 15;161(12):6455–6458. [PubMed] [Google Scholar]
  42. Shapiro V. S., Truitt K. E., Imboden J. B., Weiss A. CD28 mediates transcriptional upregulation of the interleukin-2 (IL-2) promoter through a composite element containing the CD28RE and NF-IL-2B AP-1 sites. Mol Cell Biol. 1997 Jul;17(7):4051–4058. doi: 10.1128/mcb.17.7.4051. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Song H. Y., Régnier C. H., Kirschning C. J., Goeddel D. V., Rothe M. Tumor necrosis factor (TNF)-mediated kinase cascades: bifurcation of nuclear factor-kappaB and c-jun N-terminal kinase (JNK/SAPK) pathways at TNF receptor-associated factor 2. Proc Natl Acad Sci U S A. 1997 Sep 2;94(18):9792–9796. doi: 10.1073/pnas.94.18.9792. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Su B., Jacinto E., Hibi M., Kallunki T., Karin M., Ben-Neriah Y. JNK is involved in signal integration during costimulation of T lymphocytes. Cell. 1994 Jun 3;77(5):727–736. doi: 10.1016/0092-8674(94)90056-6. [DOI] [PubMed] [Google Scholar]
  45. Tassi E., Biesova Z., Di Fiore P. P., Gutkind J. S., Wong W. T. Human JIK, a novel member of the STE20 kinase family that inhibits JNK and is negatively regulated by epidermal growth factor. J Biol Chem. 1999 Nov 19;274(47):33287–33295. doi: 10.1074/jbc.274.47.33287. [DOI] [PubMed] [Google Scholar]
  46. Truitt K. E., Hicks C. M., Imboden J. B. Stimulation of CD28 triggers an association between CD28 and phosphatidylinositol 3-kinase in Jurkat T cells. J Exp Med. 1994 Mar 1;179(3):1071–1076. doi: 10.1084/jem.179.3.1071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Umlauf S. W., Beverly B., Kang S. M., Brorson K., Tran A. C., Schwartz R. H. Molecular regulation of the IL-2 gene: rheostatic control of the immune system. Immunol Rev. 1993 Jun;133:177–197. doi: 10.1111/j.1600-065x.1993.tb01516.x. [DOI] [PubMed] [Google Scholar]
  48. Weiss A., Littman D. R. Signal transduction by lymphocyte antigen receptors. Cell. 1994 Jan 28;76(2):263–274. doi: 10.1016/0092-8674(94)90334-4. [DOI] [PubMed] [Google Scholar]
  49. Whitmarsh A. J., Davis R. J. Transcription factor AP-1 regulation by mitogen-activated protein kinase signal transduction pathways. J Mol Med (Berl) 1996 Oct;74(10):589–607. doi: 10.1007/s001090050063. [DOI] [PubMed] [Google Scholar]
  50. Yujiri T., Sather S., Fanger G. R., Johnson G. L. Role of MEKK1 in cell survival and activation of JNK and ERK pathways defined by targeted gene disruption. Science. 1998 Dec 4;282(5395):1911–1914. doi: 10.1126/science.282.5395.1911. [DOI] [PubMed] [Google Scholar]
  51. Zandi E., Rothwarf D. M., Delhase M., Hayakawa M., Karin M. The IkappaB kinase complex (IKK) contains two kinase subunits, IKKalpha and IKKbeta, necessary for IkappaB phosphorylation and NF-kappaB activation. Cell. 1997 Oct 17;91(2):243–252. doi: 10.1016/s0092-8674(00)80406-7. [DOI] [PubMed] [Google Scholar]

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