Skip to main content
NCBI home page
The EMBO Journal logoLink to The EMBO Journal
. 1996 May 15;15(10):2425–2433.

Suppression of interleukin-3-induced gene expression by a C-terminal truncated Stat5: role of Stat5 in proliferation.

A L Mui 1, H Wakao 1, T Kinoshita 1, T Kitamura 1, A Miyajima 1
PMCID: PMC450174  PMID: 8665850

Abstract

Interleukin-3 (IL3) was shown recently to utilize the transcription factor Stat5, but the genes regulated by this pathway and the biological consequence of Stat5 activation remained to be determined. In order to study the role of Stat5 in IL3 signalling, we constructed a dominant-negative Stat5 protein by C-terminal truncation, and inducibly expressed it in an IL3-dependent cell line. The effect of dominant-negative Stat5 induction on expression of IL3 early response genes was examined, and expression of several genes, including cis, osm and pim-1 was inhibited profoundly. The expression of c-fos was also reduced, but to a lesser extent. While activated Ras alone (though not Stat5 alone) could induce c-fos, maximal expression required the action of both Ras and Stat5. Interestingly, although the membrane-proximal region of the IL3 receptor beta-chain is responsible for both Jak2-Stat5 activation and c-myc induction, c-myc levels were not affected by the dominant-negative Stat5. Thus, the signals directed by this membrane-proximal domain, which is essential for transducing a DNA synthesis signal, can be separated further into Stat5 or c-myc pathways. The net effect of dominant-negative Stat5 expression was partial inhibition of IL3-dependent growth. This provides the first direct evidence that Stat5 is involved in regulation of cell proliferation.

Full text

PDF
2425

Images in this article

2427Image
on p.2427
2427Image
on p.2427
2428Image
on p.2428
2429Image
on p.2429
2430Image
on p.2430

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arai K. I., Lee F., Miyajima A., Miyatake S., Arai N., Yokota T. Cytokines: coordinators of immune and inflammatory responses. Annu Rev Biochem. 1990;59:783–836. doi: 10.1146/annurev.bi.59.070190.004031. [DOI] [PubMed] [Google Scholar]
  2. Azam M., Erdjument-Bromage H., Kreider B. L., Xia M., Quelle F., Basu R., Saris C., Tempst P., Ihle J. N., Schindler C. Interleukin-3 signals through multiple isoforms of Stat5. EMBO J. 1995 Apr 3;14(7):1402–1411. doi: 10.1002/j.1460-2075.1995.tb07126.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Barahmand-pour F., Meinke A., Eilers A., Gouilleux F., Groner B., Decker T. Colony-stimulating factors and interferon-gamma activate a protein related to MGF-Stat 5 to cause formation of the differentiation-induced factor in myeloid cells. FEBS Lett. 1995 Feb 20;360(1):29–33. doi: 10.1016/0014-5793(95)00072-h. [DOI] [PubMed] [Google Scholar]
  4. Bluyssen H. A., Muzaffar R., Vlieststra R. J., van der Made A. C., Leung S., Stark G. R., Kerr I. M., Trapman J., Levy D. E. Combinatorial association and abundance of components of interferon-stimulated gene factor 3 dictate the selectivity of interferon responses. Proc Natl Acad Sci U S A. 1995 Jun 6;92(12):5645–5649. doi: 10.1073/pnas.92.12.5645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Damen J. E., Wakao H., Miyajima A., Krosl J., Humphries R. K., Cutler R. L., Krystal G. Tyrosine 343 in the erythropoietin receptor positively regulates erythropoietin-induced cell proliferation and Stat5 activation. EMBO J. 1995 Nov 15;14(22):5557–5568. doi: 10.1002/j.1460-2075.1995.tb00243.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Danial N. N., Pernis A., Rothman P. B. Jak-STAT signaling induced by the v-abl oncogene. Science. 1995 Sep 29;269(5232):1875–1877. doi: 10.1126/science.7569929. [DOI] [PubMed] [Google Scholar]
  7. Darnell J. E., Jr, Kerr I. M., Stark G. R. Jak-STAT pathways and transcriptional activation in response to IFNs and other extracellular signaling proteins. Science. 1994 Jun 3;264(5164):1415–1421. doi: 10.1126/science.8197455. [DOI] [PubMed] [Google Scholar]
  8. Domen J., van der Lugt N. M., Laird P. W., Saris C. J., Clarke A. R., Hooper M. L., Berns A. Impaired interleukin-3 response in Pim-1-deficient bone marrow-derived mast cells. Blood. 1993 Sep 1;82(5):1445–1452. [PubMed] [Google Scholar]
  9. Eilers A., Georgellis D., Klose B., Schindler C., Ziemiecki A., Harpur A. G., Wilks A. F., Decker T. Differentiation-regulated serine phosphorylation of STAT1 promotes GAF activation in macrophages. Mol Cell Biol. 1995 Jul;15(7):3579–3586. doi: 10.1128/mcb.15.7.3579. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Friedmann M. C., Migone T. S., Russell S. M., Leonard W. J. Different interleukin 2 receptor beta-chain tyrosines couple to at least two signaling pathways and synergistically mediate interleukin 2-induced proliferation. Proc Natl Acad Sci U S A. 1996 Mar 5;93(5):2077–2082. doi: 10.1073/pnas.93.5.2077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fujii H., Nakagawa Y., Schindler U., Kawahara A., Mori H., Gouilleux F., Groner B., Ihle J. N., Minami Y., Miyazaki T. Activation of Stat5 by interleukin 2 requires a carboxyl-terminal region of the interleukin 2 receptor beta chain but is not essential for the proliferative signal transmission. Proc Natl Acad Sci U S A. 1995 Jun 6;92(12):5482–5486. doi: 10.1073/pnas.92.12.5482. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gossen M., Bujard H. Tight control of gene expression in mammalian cells by tetracycline-responsive promoters. Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5547–5551. doi: 10.1073/pnas.89.12.5547. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gouilleux F., Pallard C., Dusanter-Fourt I., Wakao H., Haldosen L. A., Norstedt G., Levy D., Groner B. Prolactin, growth hormone, erythropoietin and granulocyte-macrophage colony stimulating factor induce MGF-Stat5 DNA binding activity. EMBO J. 1995 May 1;14(9):2005–2013. doi: 10.1002/j.1460-2075.1995.tb07192.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gouilleux F., Wakao H., Mundt M., Groner B. Prolactin induces phosphorylation of Tyr694 of Stat5 (MGF), a prerequisite for DNA binding and induction of transcription. EMBO J. 1994 Sep 15;13(18):4361–4369. doi: 10.1002/j.1460-2075.1994.tb06756.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hara E., Yamaguchi T., Nojima H., Ide T., Campisi J., Okayama H., Oda K. Id-related genes encoding helix-loop-helix proteins are required for G1 progression and are repressed in senescent human fibroblasts. J Biol Chem. 1994 Jan 21;269(3):2139–2145. [PubMed] [Google Scholar]
  16. Ho A. S., Wei S. H., Mui A. L., Miyajima A., Moore K. W. Functional regions of the mouse interleukin-10 receptor cytoplasmic domain. Mol Cell Biol. 1995 Sep;15(9):5043–5053. doi: 10.1128/mcb.15.9.5043. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hou J., Schindler U., Henzel W. J., Ho T. C., Brasseur M., McKnight S. L. An interleukin-4-induced transcription factor: IL-4 Stat. Science. 1994 Sep 16;265(5179):1701–1706. doi: 10.1126/science.8085155. [DOI] [PubMed] [Google Scholar]
  18. Ihle J. N., Witthuhn B. A., Quelle F. W., Yamamoto K., Thierfelder W. E., Kreider B., Silvennoinen O. Signaling by the cytokine receptor superfamily: JAKs and STATs. Trends Biochem Sci. 1994 May;19(5):222–227. doi: 10.1016/0968-0004(94)90026-4. [DOI] [PubMed] [Google Scholar]
  19. Kinoshita T., Yokota T., Arai K., Miyajima A. Regulation of Bcl-2 expression by oncogenic Ras protein in hematopoietic cells. Oncogene. 1995 Jun 1;10(11):2207–2212. [PubMed] [Google Scholar]
  20. Kinoshita T., Yokota T., Arai K., Miyajima A. Suppression of apoptotic death in hematopoietic cells by signalling through the IL-3/GM-CSF receptors. EMBO J. 1995 Jan 16;14(2):266–275. doi: 10.1002/j.1460-2075.1995.tb07000.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lilly M., Le T., Holland P., Hendrickson S. L. Sustained expression of the pim-1 kinase is specifically induced in myeloid cells by cytokines whose receptors are structurally related. Oncogene. 1992 Apr;7(4):727–732. [PubMed] [Google Scholar]
  22. Lin J. X., Migone T. S., Tsang M., Friedmann M., Weatherbee J. A., Zhou L., Yamauchi A., Bloom E. T., Mietz J., John S. The role of shared receptor motifs and common Stat proteins in the generation of cytokine pleiotropy and redundancy by IL-2, IL-4, IL-7, IL-13, and IL-15. Immunity. 1995 Apr;2(4):331–339. doi: 10.1016/1074-7613(95)90141-8. [DOI] [PubMed] [Google Scholar]
  23. Migone T. S., Lin J. X., Cereseto A., Mulloy J. C., O'Shea J. J., Franchini G., Leonard W. J. Constitutively activated Jak-STAT pathway in T cells transformed with HTLV-I. Science. 1995 Jul 7;269(5220):79–81. doi: 10.1126/science.7604283. [DOI] [PubMed] [Google Scholar]
  24. Miyajima A., Kitamura T., Harada N., Yokota T., Arai K. Cytokine receptors and signal transduction. Annu Rev Immunol. 1992;10:295–331. doi: 10.1146/annurev.iy.10.040192.001455. [DOI] [PubMed] [Google Scholar]
  25. Miyajima A., Mui A. L., Ogorochi T., Sakamaki K. Receptors for granulocyte-macrophage colony-stimulating factor, interleukin-3, and interleukin-5. Blood. 1993 Oct 1;82(7):1960–1974. [PubMed] [Google Scholar]
  26. Mui A. L., Wakao H., O'Farrell A. M., Harada N., Miyajima A. Interleukin-3, granulocyte-macrophage colony stimulating factor and interleukin-5 transduce signals through two STAT5 homologs. EMBO J. 1995 Mar 15;14(6):1166–1175. doi: 10.1002/j.1460-2075.1995.tb07100.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Quelle F. W., Sato N., Witthuhn B. A., Inhorn R. C., Eder M., Miyajima A., Griffin J. D., Ihle J. N. JAK2 associates with the beta c chain of the receptor for granulocyte-macrophage colony-stimulating factor, and its activation requires the membrane-proximal region. Mol Cell Biol. 1994 Jul;14(7):4335–4341. doi: 10.1128/mcb.14.7.4335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Quelle F. W., Shimoda K., Thierfelder W., Fischer C., Kim A., Ruben S. M., Cleveland J. L., Pierce J. H., Keegan A. D., Nelms K. Cloning of murine Stat6 and human Stat6, Stat proteins that are tyrosine phosphorylated in responses to IL-4 and IL-3 but are not required for mitogenesis. Mol Cell Biol. 1995 Jun;15(6):3336–3343. doi: 10.1128/mcb.15.6.3336. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Renz M., Neuberg M., Kurz C., Bravo R., Müller R. Regulation of c-fos transcription in mouse fibroblasts: identification of DNase I-hypersensitive sites and regulatory upstream sequences. EMBO J. 1985 Dec 30;4(13B):3711–3716. doi: 10.1002/j.1460-2075.1985.tb04139.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sakamaki K., Miyajima I., Kitamura T., Miyajima A. Critical cytoplasmic domains of the common beta subunit of the human GM-CSF, IL-3 and IL-5 receptors for growth signal transduction and tyrosine phosphorylation. EMBO J. 1992 Oct;11(10):3541–3549. doi: 10.1002/j.1460-2075.1992.tb05437.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Samuels M. L., Weber M. J., Bishop J. M., McMahon M. Conditional transformation of cells and rapid activation of the mitogen-activated protein kinase cascade by an estradiol-dependent human raf-1 protein kinase. Mol Cell Biol. 1993 Oct;13(10):6241–6252. doi: 10.1128/mcb.13.10.6241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Sato N., Sakamaki K., Terada N., Arai K., Miyajima A. Signal transduction by the high-affinity GM-CSF receptor: two distinct cytoplasmic regions of the common beta subunit responsible for different signaling. EMBO J. 1993 Nov;12(11):4181–4189. doi: 10.1002/j.1460-2075.1993.tb06102.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Schmitt-Ney M., Doppler W., Ball R. K., Groner B. Beta-casein gene promoter activity is regulated by the hormone-mediated relief of transcriptional repression and a mammary-gland-specific nuclear factor. Mol Cell Biol. 1991 Jul;11(7):3745–3755. doi: 10.1128/mcb.11.7.3745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Treisman R. Identification of a protein-binding site that mediates transcriptional response of the c-fos gene to serum factors. Cell. 1986 Aug 15;46(4):567–574. doi: 10.1016/0092-8674(86)90882-2. [DOI] [PubMed] [Google Scholar]
  35. Voronova A. F., Lee F. The E2A and tal-1 helix-loop-helix proteins associate in vivo and are modulated by Id proteins during interleukin 6-induced myeloid differentiation. Proc Natl Acad Sci U S A. 1994 Jun 21;91(13):5952–5956. doi: 10.1073/pnas.91.13.5952. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Wakao H., Gouilleux F., Groner B. Mammary gland factor (MGF) is a novel member of the cytokine regulated transcription factor gene family and confers the prolactin response. EMBO J. 1994 May 1;13(9):2182–2191. doi: 10.1002/j.1460-2075.1994.tb06495.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Wakao H., Harada N., Kitamura T., Mui A. L., Miyajima A. Interleukin 2 and erythropoietin activate STAT5/MGF via distinct pathways. EMBO J. 1995 Jun 1;14(11):2527–2535. doi: 10.1002/j.1460-2075.1995.tb07250.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Waxman D. J., Ram P. A., Park S. H., Choi H. K. Intermittent plasma growth hormone triggers tyrosine phosphorylation and nuclear translocation of a liver-expressed, Stat 5-related DNA binding protein. Proposed role as an intracellular regulator of male-specific liver gene transcription. J Biol Chem. 1995 Jun 2;270(22):13262–13270. doi: 10.1074/jbc.270.22.13262. [DOI] [PubMed] [Google Scholar]
  39. Yip-Schneider M. T., Horie M., Broxmeyer H. E. Transcriptional induction of pim-1 protein kinase gene expression by interferon gamma and posttranscriptional effects on costimulation with steel factor. Blood. 1995 Jun 15;85(12):3494–3502. [PubMed] [Google Scholar]
  40. Yoshimura A., Ichihara M., Kinjyo I., Moriyama M., Copeland N. G., Gilbert D. J., Jenkins N. A., Hara T., Miyajima A. Mouse oncostatin M: an immediate early gene induced by multiple cytokines through the JAK-STAT5 pathway. EMBO J. 1996 Mar 1;15(5):1055–1063. [PMC free article] [PubMed] [Google Scholar]
  41. Yoshimura A., Ohkubo T., Kiguchi T., Jenkins N. A., Gilbert D. J., Copeland N. G., Hara T., Miyajima A. A novel cytokine-inducible gene CIS encodes an SH2-containing protein that binds to tyrosine-phosphorylated interleukin 3 and erythropoietin receptors. EMBO J. 1995 Jun 15;14(12):2816–2826. doi: 10.1002/j.1460-2075.1995.tb07281.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Yu C. L., Meyer D. J., Campbell G. S., Larner A. C., Carter-Su C., Schwartz J., Jove R. Enhanced DNA-binding activity of a Stat3-related protein in cells transformed by the Src oncoprotein. Science. 1995 Jul 7;269(5220):81–83. doi: 10.1126/science.7541555. [DOI] [PubMed] [Google Scholar]
  43. Zhang X., Blenis J., Li H. C., Schindler C., Chen-Kiang S. Requirement of serine phosphorylation for formation of STAT-promoter complexes. Science. 1995 Mar 31;267(5206):1990–1994. doi: 10.1126/science.7701321. [DOI] [PubMed] [Google Scholar]
  44. van Lohuizen M., Verbeek S., Krimpenfort P., Domen J., Saris C., Radaszkiewicz T., Berns A. Predisposition to lymphomagenesis in pim-1 transgenic mice: cooperation with c-myc and N-myc in murine leukemia virus-induced tumors. Cell. 1989 Feb 24;56(4):673–682. doi: 10.1016/0092-8674(89)90589-8. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES