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. 1992 May 1;89(9):4052–4056. doi: 10.1073/pnas.89.9.4052

A single gene encodes two isoforms of the p70 S6 kinase: activation upon mitogenic stimulation.

C Reinhard 1, G Thomas 1, S C Kozma 1
PMCID: PMC525630  PMID: 1570332

Abstract

Previously, two cDNA clones were isolated from a rat liver or hepatoma cDNA library for the mitogenic-activated p70 S6 kinase (p70s6k). Except for a single amino acid change and a 23-amino acid N-terminal extension in the latter clone, the open reading frames of the two clones are identical. A probe common to both clones also revealed four distinct transcripts. Here, by using specific probes, it was possible to show which transcript corresponds to which clone and that both clones are derived from the same gene. Furthermore, analysis of in vitro translation products using specific antibodies demonstrates that both clones encode the p70s6k but that the clone harboring the 23-amino acid extension also encodes an additional isoform of the kinase, referred to as p85s6k. It could be shown by using the specific antibody to the p85s6k that this isoform of the kinase is present in rat liver and is activated after mitogenic stimulation of quiescent Swiss 3T3 cells.

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Selected References

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

  1. Ballou L. M., Luther H., Thomas G. MAP2 kinase and 70K S6 kinase lie on distinct signalling pathways. Nature. 1991 Jan 24;349(6307):348–350. doi: 10.1038/349348a0. [DOI] [PubMed] [Google Scholar]
  2. Ballou L. M., Siegmann M., Thomas G. S6 kinase in quiescent Swiss mouse 3T3 cells is activated by phosphorylation in response to serum treatment. Proc Natl Acad Sci U S A. 1988 Oct;85(19):7154–7158. doi: 10.1073/pnas.85.19.7154. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Banerjee P., Ahmad M. F., Grove J. R., Kozlosky C., Price D. J., Avruch J. Molecular structure of a major insulin/mitogen-activated 70-kDa S6 protein kinase. Proc Natl Acad Sci U S A. 1990 Nov;87(21):8550–8554. doi: 10.1073/pnas.87.21.8550. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Blenis J., Chung J., Erikson E., Alcorta D. A., Erikson R. L. Distinct mechanisms for the activation of the RSK kinases/MAP2 kinase/pp90rsk and pp70-S6 kinase signaling systems are indicated by inhibition of protein synthesis. Cell Growth Differ. 1991 Jun;2(6):279–285. [PubMed] [Google Scholar]
  5. Chen R. H., Blenis J. Identification of Xenopus S6 protein kinase homologs (pp90rsk) in somatic cells: phosphorylation and activation during initiation of cell proliferation. Mol Cell Biol. 1990 Jun;10(6):3204–3215. doi: 10.1128/mcb.10.6.3204. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  7. Cochrane A. W., Perkins A., Rosen C. A. Identification of sequences important in the nucleolar localization of human immunodeficiency virus Rev: relevance of nucleolar localization to function. J Virol. 1990 Feb;64(2):881–885. doi: 10.1128/jvi.64.2.881-885.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Erikson R. L. Structure, expression, and regulation of protein kinases involved in the phosphorylation of ribosomal protein S6. J Biol Chem. 1991 Apr 5;266(10):6007–6010. [PubMed] [Google Scholar]
  9. Ferrari S., Bandi H. R., Hofsteenge J., Bussian B. M., Thomas G. Mitogen-activated 70K S6 kinase. Identification of in vitro 40 S ribosomal S6 phosphorylation sites. J Biol Chem. 1991 Nov 25;266(33):22770–22775. [PubMed] [Google Scholar]
  10. Flotow H., Thomas G. Substrate recognition determinants of the mitogen-activated 70K S6 kinase from rat liver. J Biol Chem. 1992 Feb 15;267(5):3074–3078. [PubMed] [Google Scholar]
  11. Franco R., Rosenfeld M. G. Hormonally inducible phosphorylation of a nuclear pool of ribosomal protein S6. J Biol Chem. 1990 Mar 15;265(8):4321–4325. [PubMed] [Google Scholar]
  12. Garcia-Bustos J., Heitman J., Hall M. N. Nuclear protein localization. Biochim Biophys Acta. 1991 Mar 7;1071(1):83–101. doi: 10.1016/0304-4157(91)90013-m. [DOI] [PubMed] [Google Scholar]
  13. Grove J. R., Banerjee P., Balasubramanyam A., Coffer P. J., Price D. J., Avruch J., Woodgett J. R. Cloning and expression of two human p70 S6 kinase polypeptides differing only at their amino termini. Mol Cell Biol. 1991 Nov;11(11):5541–5550. doi: 10.1128/mcb.11.11.5541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Harmann B., Kilimann M. W. cDNA encoding a 59 kDa homolog of ribosomal protein S6 kinase from rabbit liver. FEBS Lett. 1990 Oct 29;273(1-2):248–252. doi: 10.1016/0014-5793(90)81096-7. [DOI] [PubMed] [Google Scholar]
  15. Hope T. J., Huang X. J., McDonald D., Parslow T. G. Steroid-receptor fusion of the human immunodeficiency virus type 1 Rev transactivator: mapping cryptic functions of the arginine-rich motif. Proc Natl Acad Sci U S A. 1990 Oct;87(19):7787–7791. doi: 10.1073/pnas.87.19.7787. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jackson R. J., Hunt T. Preparation and use of nuclease-treated rabbit reticulocyte lysates for the translation of eukaryotic messenger RNA. Methods Enzymol. 1983;96:50–74. doi: 10.1016/s0076-6879(83)96008-1. [DOI] [PubMed] [Google Scholar]
  17. Jenö P., Ballou L. M., Novak-Hofer I., Thomas G. Identification and characterization of a mitogen-activated S6 kinase. Proc Natl Acad Sci U S A. 1988 Jan;85(2):406–410. doi: 10.1073/pnas.85.2.406. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Jenö P., Jäggi N., Luther H., Siegmann M., Thomas G. Purification and characterization of a 40 S ribosomal protein S6 kinase from vanadate-stimulated Swiss 3T3 cells. J Biol Chem. 1989 Jan 15;264(2):1293–1297. [PubMed] [Google Scholar]
  19. Kozak M. Structural features in eukaryotic mRNAs that modulate the initiation of translation. J Biol Chem. 1991 Oct 25;266(30):19867–19870. [PubMed] [Google Scholar]
  20. Kozma S. C., Ferrari S., Bassand P., Siegmann M., Totty N., Thomas G. Cloning of the mitogen-activated S6 kinase from rat liver reveals an enzyme of the second messenger subfamily. Proc Natl Acad Sci U S A. 1990 Oct;87(19):7365–7369. doi: 10.1073/pnas.87.19.7365. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kozma S. C., Lane H. A., Ferrari S., Luther H., Siegmann M., Thomas G. A stimulated S6 kinase from rat liver: identity with the mitogen activated S6 kinase of 3T3 cells. EMBO J. 1989 Dec 20;8(13):4125–4132. doi: 10.1002/j.1460-2075.1989.tb08597.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Krieg J., Hofsteenge J., Thomas G. Identification of the 40 S ribosomal protein S6 phosphorylation sites induced by cycloheximide. J Biol Chem. 1988 Aug 15;263(23):11473–11477. [PubMed] [Google Scholar]
  23. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  24. Lane H. A., Morley S. J., Dorée M., Kozma S. C., Thomas G. Identification and early activation of a Xenopus laevis p70s6k following progesterone-induced meiotic maturation. EMBO J. 1992 May;11(5):1743–1749. doi: 10.1002/j.1460-2075.1992.tb05226.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lane H. A., Thomas G. Purification and properties of mitogen-activated S6 kinase from rat liver and 3T3 cells. Methods Enzymol. 1991;200:268–291. doi: 10.1016/0076-6879(91)00146-n. [DOI] [PubMed] [Google Scholar]
  26. Mahadevan L. C., Edwards D. R. Signalling and superinduction. Nature. 1991 Feb 28;349(6312):747–748. doi: 10.1038/349747c0. [DOI] [PubMed] [Google Scholar]
  27. Olivier A. R., Thomas G. Three forms of phosphatase type 1 in Swiss 3T3 fibroblasts. Free catalytic subunit appears to mediate s6 dephosphorylation. J Biol Chem. 1990 Dec 25;265(36):22460–22466. [PubMed] [Google Scholar]
  28. Price D. J., Nemenoff R. A., Avruch J. Purification of a hepatic S6 kinase from cycloheximide-treated Rats. J Biol Chem. 1989 Aug 15;264(23):13825–13833. [PubMed] [Google Scholar]
  29. Rihs H. P., Jans D. A., Fan H., Peters R. The rate of nuclear cytoplasmic protein transport is determined by the casein kinase II site flanking the nuclear localization sequence of the SV40 T-antigen. EMBO J. 1991 Mar;10(3):633–639. doi: 10.1002/j.1460-2075.1991.tb07991.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Schowalter D. B., Sommer S. S. The generation of radiolabeled DNA and RNA probes with polymerase chain reaction. Anal Biochem. 1989 Feb 15;177(1):90–94. doi: 10.1016/0003-2697(89)90019-5. [DOI] [PubMed] [Google Scholar]
  31. Sturgill T. W., Wu J. Recent progress in characterization of protein kinase cascades for phosphorylation of ribosomal protein S6. Biochim Biophys Acta. 1991 May 17;1092(3):350–357. doi: 10.1016/s0167-4889(97)90012-4. [DOI] [PubMed] [Google Scholar]
  32. Susa M., Olivier A. R., Fabbro D., Thomas G. EGF induces biphasic S6 kinase activation: late phase is protein kinase C-dependent and contributes to mitogenicity. Cell. 1989 Jun 2;57(5):817–824. doi: 10.1016/0092-8674(89)90796-4. [DOI] [PubMed] [Google Scholar]
  33. Susa M., Thomas G. Identical Mr 70,000 S6 kinase is activated biphasically by epidermal growth factor: a phosphopeptide that characterizes the late phase. Proc Natl Acad Sci U S A. 1990 Sep;87(18):7040–7044. doi: 10.1073/pnas.87.18.7040. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Thomas G. MAP kinase by any other name smells just as sweet. Cell. 1992 Jan 10;68(1):3–6. doi: 10.1016/0092-8674(92)90199-m. [DOI] [PubMed] [Google Scholar]

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