Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1996 Dec 15;24(24):4969–4977. doi: 10.1093/nar/24.24.4969

In vivo generation of 3' and 5' truncated species in the process of c-myc mRNA decay.

P Ioannidis 1, M Havredaki 1, N Courtis 1, T Trangas 1
PMCID: PMC146348  PMID: 9016668

Abstract

It has been demonstrated that the half-life of c-myc mRNA is modulated in response to physiological agents. The elucidation of the decay process and the identification of the critical steps in the in vivo c-myc mRNA degradation pathway can be approached by following the fate of c-myc mRNA under the influence of such factors. IFN-alpha was the factor used to modulate c-myc mRNA half-life in HeLa 1C5 cells, a stable clone derived from HeLa cells. This cell line carries multiple copies of the c-myc gene, under the control of the dexamethasone inducible mouse mammary tumor virus-long terminal repeat (MMTV-LTR). Exposure of HeLa 1C5 cells to IFN-alpha resulted in a further 2-fold increase over the dexamethasone-induced c-myc mRNA. However, the c-myc mRNA in IFN-alpha treated cells was less stable than that in the control cells. RNase H mapping of the 3' untranslated region of c-myc mRNA revealed, in addition to the full length mRNA, three smaller fragments. These fragments were proven to be truncated, non-adenylated c-myc mRNA species generated in vivo. Exposure of HeLa 1C5 cells to Interferon-alpha before induction with dexamethasone resulted in the enhanced presence of these intermediates. RNase H analysis of c-myc mRNA after actinomycin D chase revealed that deadenylation led to the formation of a relatively more stable oligoadenylated c-myc mRNA population which did not appear to be precursor to the truncated intermediates. The detection of truncated 3' end c-myc mRNA adenylated fragments as well, implies that the c-myc mRNA degradation process may follow an alternative pathway possibly involving endonucleolytic cleavage.

Full Text

The Full Text of this article is available as a PDF (279.9 KB).

Selected References

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

  1. Alberta J. A., Rundell K., Stiles C. D. Identification of an activity that interacts with the 3'-untranslated region of c-myc mRNA and the role of its target sequence in mediating rapid mRNA degradation. J Biol Chem. 1994 Feb 11;269(6):4532–4538. [PubMed] [Google Scholar]
  2. Bernstein P. L., Herrick D. J., Prokipcak R. D., Ross J. Control of c-myc mRNA half-life in vitro by a protein capable of binding to a coding region stability determinant. Genes Dev. 1992 Apr;6(4):642–654. doi: 10.1101/gad.6.4.642. [DOI] [PubMed] [Google Scholar]
  3. Binder R., Horowitz J. A., Basilion J. P., Koeller D. M., Klausner R. D., Harford J. B. Evidence that the pathway of transferrin receptor mRNA degradation involves an endonucleolytic cleavage within the 3' UTR and does not involve poly(A) tail shortening. EMBO J. 1994 Apr 15;13(8):1969–1980. doi: 10.1002/j.1460-2075.1994.tb06466.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Binder R., Hwang S. P., Ratnasabapathy R., Williams D. L. Degradation of apolipoprotein II mRNA occurs via endonucleolytic cleavage at 5'-AAU-3'/5'-UAA-3' elements in single-stranded loop domains of the 3'-noncoding region. J Biol Chem. 1989 Oct 5;264(28):16910–16918. [PubMed] [Google Scholar]
  5. Bissonnette R. P., Echeverri F., Mahboubi A., Green D. R. Apoptotic cell death induced by c-myc is inhibited by bcl-2. Nature. 1992 Oct 8;359(6395):552–554. doi: 10.1038/359552a0. [DOI] [PubMed] [Google Scholar]
  6. Blanchard J. M., Piechaczyk M., Dani C., Chambard J. C., Franchi A., Pouyssegur J., Jeanteur P. c-myc gene is transcribed at high rate in G0-arrested fibroblasts and is post-transcriptionally regulated in response to growth factors. Nature. 1985 Oct 3;317(6036):443–445. doi: 10.1038/317443a0. [DOI] [PubMed] [Google Scholar]
  7. Bonnieu A., Roux P., Marty L., Jeanteur P., Piechaczyk M. AUUUA motifs are dispensable for rapid degradation of the mouse c-myc RNA. Oncogene. 1990 Oct;5(10):1585–1588. [PubMed] [Google Scholar]
  8. Brewer G. An A + U-rich element RNA-binding factor regulates c-myc mRNA stability in vitro. Mol Cell Biol. 1991 May;11(5):2460–2466. doi: 10.1128/mcb.11.5.2460. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Brewer G., Ross J. Messenger RNA turnover in cell-free extracts. Methods Enzymol. 1990;181:202–209. doi: 10.1016/0076-6879(90)81122-b. [DOI] [PubMed] [Google Scholar]
  10. Brewer G., Ross J. Poly(A) shortening and degradation of the 3' A+U-rich sequences of human c-myc mRNA in a cell-free system. Mol Cell Biol. 1988 Apr;8(4):1697–1708. doi: 10.1128/mcb.8.4.1697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Chatterjee D., Savarese T. M. Posttranscriptional regulation of c-myc proto-oncogene expression and growth inhibition by recombinant human interferon-beta ser17 in a human colon carcinoma cell line. Cancer Chemother Pharmacol. 1992;30(1):12–20. doi: 10.1007/BF00686479. [DOI] [PubMed] [Google Scholar]
  12. Chen C. Y., Shyu A. B. Selective degradation of early-response-gene mRNAs: functional analyses of sequence features of the AU-rich elements. Mol Cell Biol. 1994 Dec;14(12):8471–8482. doi: 10.1128/mcb.14.12.8471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. 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]
  14. Dani C., Blanchard J. M., Piechaczyk M., El Sabouty S., Marty L., Jeanteur P. Extreme instability of myc mRNA in normal and transformed human cells. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7046–7050. doi: 10.1073/pnas.81.22.7046. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Dani C., Mechti N., Piechaczyk M., Lebleu B., Jeanteur P., Blanchard J. M. Increased rate of degradation of c-myc mRNA in interferon-treated Daudi cells. Proc Natl Acad Sci U S A. 1985 Aug;82(15):4896–4899. doi: 10.1073/pnas.82.15.4896. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gazin C., Dupont de Dinechin S., Hampe A., Masson J. M., Martin P., Stehelin D., Galibert F. Nucleotide sequence of the human c-myc locus: provocative open reading frame within the first exon. EMBO J. 1984 Feb;3(2):383–387. doi: 10.1002/j.1460-2075.1984.tb01816.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jones T. R., Cole M. D. Rapid cytoplasmic turnover of c-myc mRNA: requirement of the 3' untranslated sequences. Mol Cell Biol. 1987 Dec;7(12):4513–4521. doi: 10.1128/mcb.7.12.4513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kelly J. M., Gilbert C. S., Stark G. R., Kerr I. M. Differential regulation of interferon-induced mRNAs and c-myc mRNA by alpha- and gamma-interferons. Eur J Biochem. 1985 Dec 2;153(2):367–371. doi: 10.1111/j.1432-1033.1985.tb09312.x. [DOI] [PubMed] [Google Scholar]
  19. Kerr I. M. The 2-5A system: a personal view. J Interferon Res. 1987 Oct;7(5):505–510. doi: 10.1089/jir.1987.7.505. [DOI] [PubMed] [Google Scholar]
  20. Kimchi A. Autocrine interferon and the suppression of the c-myc nuclear oncogene. Interferon. 1987;8:85–110. [PubMed] [Google Scholar]
  21. Knight E., Jr, Anton E. D., Fahey D., Friedland B. K., Jonak G. J. Interferon regulates c-myc gene expression in Daudi cells at the post-transcriptional level. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1151–1154. doi: 10.1073/pnas.82.4.1151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Laird-Offringa I. A., Elfferich P., Knaken H. J., de Ruiter J., van der Eb A. J. Analysis of polyadenylation site usage of the c-myc oncogene. Nucleic Acids Res. 1989 Aug 25;17(16):6499–6514. doi: 10.1093/nar/17.16.6499. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Laird-Offringa I. A. What determines the instability of c-myc proto-oncogene mRNA? Bioessays. 1992 Feb;14(2):119–124. doi: 10.1002/bies.950140209. [DOI] [PubMed] [Google Scholar]
  24. Laird-Offringa I. A., de Wit C. L., Elfferich P., van der Eb A. J. Poly(A) tail shortening is the translation-dependent step in c-myc mRNA degradation. Mol Cell Biol. 1990 Dec;10(12):6132–6140. doi: 10.1128/mcb.10.12.6132. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Marcu K. B., Bossone S. A., Patel A. J. myc function and regulation. Annu Rev Biochem. 1992;61:809–860. doi: 10.1146/annurev.bi.61.070192.004113. [DOI] [PubMed] [Google Scholar]
  26. Mechti N., Piechaczyk M., Blanchard J. M., Marty L., Bonnieu A., Jeanteur P., Lebleu B. Transcriptional and post-transcriptional regulation of c-myc expression during the differentiation of murine erythroleukemia Friend cells. Nucleic Acids Res. 1986 Dec 22;14(24):9653–9666. doi: 10.1093/nar/14.24.9653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Meichle A., Philipp A., Eilers M. The functions of Myc proteins. Biochim Biophys Acta. 1992 Dec 16;1114(2-3):129–146. doi: 10.1016/0304-419x(92)90011-m. [DOI] [PubMed] [Google Scholar]
  28. Meurs E., Hovanessian A. G. Alpha-interferon inhibits the expression of heavy chain mu messenger RNA in Daudi cells. EMBO J. 1988 Jun;7(6):1689–1696. doi: 10.1002/j.1460-2075.1988.tb02997.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Ramsay G., Evan G. I., Bishop J. M. The protein encoded by the human proto-oncogene c-myc. Proc Natl Acad Sci U S A. 1984 Dec;81(24):7742–7746. doi: 10.1073/pnas.81.24.7742. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Ross J., Peltz S. W., Kobs G., Brewer G. Histone mRNA degradation in vivo: the first detectable step occurs at or near the 3' terminus. Mol Cell Biol. 1986 Dec;6(12):4362–4371. doi: 10.1128/mcb.6.12.4362. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Sachs A. B. Messenger RNA degradation in eukaryotes. Cell. 1993 Aug 13;74(3):413–421. doi: 10.1016/0092-8674(93)80043-e. [DOI] [PubMed] [Google Scholar]
  32. Schiavi S. C., Belasco J. G., Greenberg M. E. Regulation of proto-oncogene mRNA stability. Biochim Biophys Acta. 1992 Dec 16;1114(2-3):95–106. doi: 10.1016/0304-419x(92)90009-n. [DOI] [PubMed] [Google Scholar]
  33. Shaw G., Kamen R. A conserved AU sequence from the 3' untranslated region of GM-CSF mRNA mediates selective mRNA degradation. Cell. 1986 Aug 29;46(5):659–667. doi: 10.1016/0092-8674(86)90341-7. [DOI] [PubMed] [Google Scholar]
  34. Shi Y., Glynn J. M., Guilbert L. J., Cotter T. G., Bissonnette R. P., Green D. R. Role for c-myc in activation-induced apoptotic cell death in T cell hybridomas. Science. 1992 Jul 10;257(5067):212–214. doi: 10.1126/science.1378649. [DOI] [PubMed] [Google Scholar]
  35. Shuman S., Moss B. Vaccinia virus poly(A) polymerase. Specificity for nucleotides and nucleotide analogs. J Biol Chem. 1988 Jun 15;263(17):8405–8412. [PubMed] [Google Scholar]
  36. Shyu A. B., Belasco J. G., Greenberg M. E. Two distinct destabilizing elements in the c-fos message trigger deadenylation as a first step in rapid mRNA decay. Genes Dev. 1991 Feb;5(2):221–231. doi: 10.1101/gad.5.2.221. [DOI] [PubMed] [Google Scholar]
  37. Spencer C. A., Groudine M. Control of c-myc regulation in normal and neoplastic cells. Adv Cancer Res. 1991;56:1–48. doi: 10.1016/s0065-230x(08)60476-5. [DOI] [PubMed] [Google Scholar]
  38. Stoeckle M. Y., Guan L. Improved resolution and sensitivity of northern blots using polyacrylamide-urea gels. Biotechniques. 1993 Aug;15(2):227, 230-1. [PubMed] [Google Scholar]
  39. Stoeckle M. Y., Hanafusa H. Processing of 9E3 mRNA and regulation of its stability in normal and Rous sarcoma virus-transformed cells. Mol Cell Biol. 1989 Nov;9(11):4738–4745. doi: 10.1128/mcb.9.11.4738. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Stoeckle M. Y. Removal of a 3' non-coding sequence is an initial step in degradation of gro alpha mRNA and is regulated by interleukin-1. Nucleic Acids Res. 1992 Mar 11;20(5):1123–1127. doi: 10.1093/nar/20.5.1123. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Strobl L. J., Kohlhuber F., Mautner J., Polack A., Eick D. Absence of a paused transcription complex from the c-myc P2 promoter of the translocation chromosome in Burkitt's lymphoma cells: implication for the c-myc P1/P2 promoter shift. Oncogene. 1993 Jun;8(6):1437–1447. [PubMed] [Google Scholar]
  42. Swartwout S. G., Kinniburgh A. J. c-myc RNA degradation in growing and differentiating cells: possible alternate pathways. Mol Cell Biol. 1989 Jan;9(1):288–295. doi: 10.1128/mcb.9.1.288. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Tharun S., Sirdeshmukh R. Specific endonucleolytic cleavages of mouse albumin mRNA and their modulation during liver development. Nucleic Acids Res. 1995 Feb 25;23(4):641–646. doi: 10.1093/nar/23.4.641. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Vakalopoulou E., Schaack J., Shenk T. A 32-kilodalton protein binds to AU-rich domains in the 3' untranslated regions of rapidly degraded mRNAs. Mol Cell Biol. 1991 Jun;11(6):3355–3364. doi: 10.1128/mcb.11.6.3355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Wennborg A., Sohlberg B., Angerer D., Klein G., von Gabain A. A human RNase E-like activity that cleaves RNA sequences involved in mRNA stability control. Proc Natl Acad Sci U S A. 1995 Aug 1;92(16):7322–7326. doi: 10.1073/pnas.92.16.7322. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Wisdom R., Lee W. The protein-coding region of c-myc mRNA contains a sequence that specifies rapid mRNA turnover and induction by protein synthesis inhibitors. Genes Dev. 1991 Feb;5(2):232–243. doi: 10.1101/gad.5.2.232. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES