Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1994 Jan;14(1):207–213. doi: 10.1128/mcb.14.1.207

Identification of a thymidylate synthase ribonucleoprotein complex in human colon cancer cells.

E Chu 1, D M Voeller 1, K L Jones 1, T Takechi 1, G F Maley 1, F Maley 1, S Segal 1, C J Allegra 1
PMCID: PMC358371  PMID: 8264588

Abstract

Translation of thymidylate synthase (TS) mRNA is controlled by its own protein product, TS, in an autoregulatory manner. Direct binding of TS protein to two different cis-acting elements on the TS mRNA is associated with this translational regulation. In this study, an immunoprecipitation-reverse transcription-PCR technique was used to identify a TS ribonucleoprotein (RNP) complex in cultured human colon cancer cells. Using antibodies specific for TS protein, we show that TS is complexed in vivo with its own TS RNA. Furthermore, evidence demonstrating a direct interaction between the mRNA of the nuclear oncogene c-myc and TS protein is presented.

Full text

PDF
207

Images in this article

208Image
on p.208
209Image
on p.209
209Image
on p.209
210Image
on p.210
210Image
on p.210
211Image
on p.211

Selected References

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

  1. Ayusawa D., Shimizu K., Koyama H., Kaneda S., Takeishi K., Seno T. Cell-cycle-directed regulation of thymidylate synthase messenger RNA in human diploid fibroblasts stimulated to proliferate. J Mol Biol. 1986 Aug 20;190(4):559–567. doi: 10.1016/0022-2836(86)90241-x. [DOI] [PubMed] [Google Scholar]
  2. Bernardi A., Spahr P. F. Nucleotide sequence at the binding site for coat protein on RNA of bacteriophage R17. Proc Natl Acad Sci U S A. 1972 Oct;69(10):3033–3037. doi: 10.1073/pnas.69.10.3033. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blackwood E. M., Eisenman R. N. Max: a helix-loop-helix zipper protein that forms a sequence-specific DNA-binding complex with Myc. Science. 1991 Mar 8;251(4998):1211–1217. doi: 10.1126/science.2006410. [DOI] [PubMed] [Google Scholar]
  4. Blackwood E. M., Kretzner L., Eisenman R. N. Myc and Max function as a nucleoprotein complex. Curr Opin Genet Dev. 1992 Apr;2(2):227–235. doi: 10.1016/s0959-437x(05)80278-3. [DOI] [PubMed] [Google Scholar]
  5. Carey J., Uhlenbeck O. C. Kinetic and thermodynamic characterization of the R17 coat protein-ribonucleic acid interaction. Biochemistry. 1983 May 24;22(11):2610–2615. doi: 10.1021/bi00280a003. [DOI] [PubMed] [Google Scholar]
  6. Chu E., Drake J. C., Koeller D. M., Zinn S., Jamis-Dow C. A., Yeh G. C., Allegra C. J. Induction of thymidylate synthase associated with multidrug resistance in human breast and colon cancer cell lines. Mol Pharmacol. 1991 Feb;39(2):136–143. [PubMed] [Google Scholar]
  7. Chu E., Koeller D. M., Casey J. L., Drake J. C., Chabner B. A., Elwood P. C., Zinn S., Allegra C. J. Autoregulation of human thymidylate synthase messenger RNA translation by thymidylate synthase. Proc Natl Acad Sci U S A. 1991 Oct 15;88(20):8977–8981. doi: 10.1073/pnas.88.20.8977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chu E., Koeller D. M., Johnston P. G., Zinn S., Allegra C. J. Regulation of thymidylate synthase in human colon cancer cells treated with 5-fluorouracil and interferon-gamma. Mol Pharmacol. 1993 Apr;43(4):527–533. [PubMed] [Google Scholar]
  9. Chu E., Takimoto C. H., Voeller D., Grem J. L., Allegra C. J. Specific binding of human dihydrofolate reductase protein to dihydrofolate reductase messenger RNA in vitro. Biochemistry. 1993 May 11;32(18):4756–4760. doi: 10.1021/bi00069a009. [DOI] [PubMed] [Google Scholar]
  10. Chu E., Voeller D., Koeller D. M., Drake J. C., Takimoto C. H., Maley G. F., Maley F., Allegra C. J. Identification of an RNA binding site for human thymidylate synthase. Proc Natl Acad Sci U S A. 1993 Jan 15;90(2):517–521. doi: 10.1073/pnas.90.2.517. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cogliati T., Dunn B. K., Bar-Ner M., Cultraro C. M., Segal S. Transfected wild-type and mutant max regulate cell growth and differentiation of murine erythroleukemia cells. Oncogene. 1993 May;8(5):1263–1268. [PubMed] [Google Scholar]
  12. DePinho R. A., Schreiber-Agus N., Alt F. W. myc family oncogenes in the development of normal and neoplastic cells. Adv Cancer Res. 1991;57:1–46. doi: 10.1016/s0065-230x(08)60994-x. [DOI] [PubMed] [Google Scholar]
  13. Dolnick B. J., Zhang Z. G., Hines J. D., Rustum Y. M. Quantitation of dihydrofolate reductase and thymidylate synthase mRNAs in vivo and in vitro by polymerase chain reaction. Oncol Res. 1992;4(2):65–72. [PubMed] [Google Scholar]
  14. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  15. Haile D. J., Hentze M. W., Rouault T. A., Harford J. B., Klausner R. D. Regulation of interaction of the iron-responsive element binding protein with iron-responsive RNA elements. Mol Cell Biol. 1989 Nov;9(11):5055–5061. doi: 10.1128/mcb.9.11.5055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hentze M. W., Caughman S. W., Rouault T. A., Barriocanal J. G., Dancis A., Harford J. B., Klausner R. D. Identification of the iron-responsive element for the translational regulation of human ferritin mRNA. Science. 1987 Dec 11;238(4833):1570–1573. doi: 10.1126/science.3685996. [DOI] [PubMed] [Google Scholar]
  17. Hentze M. W., Rouault T. A., Harford J. B., Klausner R. D. Oxidation-reduction and the molecular mechanism of a regulatory RNA-protein interaction. Science. 1989 Apr 21;244(4902):357–359. doi: 10.1126/science.2711187. [DOI] [PubMed] [Google Scholar]
  18. Hershey J. W. Translational control in mammalian cells. Annu Rev Biochem. 1991;60:717–755. doi: 10.1146/annurev.bi.60.070191.003441. [DOI] [PubMed] [Google Scholar]
  19. Jenh C. H., Geyer P. K., Johnson L. F. Control of thymidylate synthase mRNA content and gene transcription in an overproducing mouse cell line. Mol Cell Biol. 1985 Oct;5(10):2527–2532. doi: 10.1128/mcb.5.10.2527. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Johnston P. G., Liang C. M., Henry S., Chabner B. A., Allegra C. J. Production and characterization of monoclonal antibodies that localize human thymidylate synthase in the cytoplasm of human cells and tissue. Cancer Res. 1991 Dec 15;51(24):6668–6676. [PubMed] [Google Scholar]
  21. Kaneda S., Nalbantoglu J., Takeishi K., Shimizu K., Gotoh O., Seno T., Ayusawa D. Structural and functional analysis of the human thymidylate synthase gene. J Biol Chem. 1990 Nov 25;265(33):20277–20284. [PubMed] [Google Scholar]
  22. Klausner R. D., Harford J. B. cis-trans models for post-transcriptional gene regulation. Science. 1989 Nov 17;246(4932):870–872. doi: 10.1126/science.2683086. [DOI] [PubMed] [Google Scholar]
  23. Kretzner L., Blackwood E. M., Eisenman R. N. Myc and Max proteins possess distinct transcriptional activities. Nature. 1992 Oct 1;359(6394):426–429. doi: 10.1038/359426a0. [DOI] [PubMed] [Google Scholar]
  24. Leibold E. A., Munro H. N. Cytoplasmic protein binds in vitro to a highly conserved sequence in the 5' untranslated region of ferritin heavy- and light-subunit mRNAs. Proc Natl Acad Sci U S A. 1988 Apr;85(7):2171–2175. doi: 10.1073/pnas.85.7.2171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lerner M. R., Boyle J. A., Hardin J. A., Steitz J. A. Two novel classes of small ribonucleoproteins detected by antibodies associated with lupus erythematosus. Science. 1981 Jan 23;211(4480):400–402. doi: 10.1126/science.6164096. [DOI] [PubMed] [Google Scholar]
  26. Lerner M. R., Steitz J. A. Antibodies to small nuclear RNAs complexed with proteins are produced by patients with systemic lupus erythematosus. Proc Natl Acad Sci U S A. 1979 Nov;76(11):5495–5499. doi: 10.1073/pnas.76.11.5495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lüscher B., Eisenman R. N. New light on Myc and Myb. Part I. Myc. Genes Dev. 1990 Dec;4(12A):2025–2035. doi: 10.1101/gad.4.12a.2025. [DOI] [PubMed] [Google Scholar]
  28. Mäkelä T. P., Koskinen P. J., Västrik I., Alitalo K. Alternative forms of Max as enhancers or suppressors of Myc-ras cotransformation. Science. 1992 Apr 17;256(5055):373–377. doi: 10.1126/science.256.5055.373. [DOI] [PubMed] [Google Scholar]
  29. Navalgund L. G., Rossana C., Muench A. J., Johnson L. F. Cell cycle regulation of thymidylate synthetase gene expression in cultured mouse fibroblasts. J Biol Chem. 1980 Aug 10;255(15):7386–7390. [PubMed] [Google Scholar]
  30. Pachter J. S., Yen T. J., Cleveland D. W. Autoregulation of tubulin expression is achieved through specific degradation of polysomal tubulin mRNAs. Cell. 1987 Oct 23;51(2):283–292. doi: 10.1016/0092-8674(87)90155-3. [DOI] [PubMed] [Google Scholar]
  31. Park J. G., Oie H. K., Sugarbaker P. H., Henslee J. G., Chen T. R., Johnson B. E., Gazdar A. Characteristics of cell lines established from human colorectal carcinoma. Cancer Res. 1987 Dec 15;47(24 Pt 1):6710–6718. [PubMed] [Google Scholar]
  32. Pestka S. The use of inhibitors in studies on protein synthesis. Methods Enzymol. 1974;30:261–282. doi: 10.1016/0076-6879(74)30030-4. [DOI] [PubMed] [Google Scholar]
  33. Steitz J. A. Immunoprecipitation of ribonucleoproteins using autoantibodies. Methods Enzymol. 1989;180:468–481. doi: 10.1016/0076-6879(89)80118-1. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES