Home LiteratureArticle Details
PMID: 8668201 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Functional mapping of the translation-dependent instability element of yeast MATalpha1 mRNA.

Molecular and cellular biology ·Vol. 16 ·No. 7 ·1996-07-00 ·Pages 3833-43

Hennigan AN, Jacobson A

Abstract

The determinants of mRNA stability include specific cis-acting destabilizing sequences located within mRNA coding and noncoding regions. We have developed an approach for mapping coding-region instability sequences in unstable yeast mRNAs that exploits the link between mRNA translation and turnover and the dependence of nonsense-mediated mRNA decay on the activity of the UPF1 gene product. This approach, which involves the systematic insertion of in-frame translational termination codons into the coding sequence of a gene of interest in a upf1delta strain, differs significantly from conventional methods for mapping cis-acting elements in that it causes minimal perturbations to overall mRNA structure. Using the previously characterized MATalpha1 mRNA as a model, we have accurately localized its 65-nucleotide instability element (IE) within the protein coding region. Termination of translation 5' to this element stabilized the MATalpha1 mRNA two- to threefold relative to wild-type transcripts. Translation through the element was sufficient to restore an unstable decay phenotype, while internal termination resulted in different extents of mRNA stabilization dependent on the precise location of ribosome stalling. Detailed mutagenesis of the element's rare-codon/AU-rich sequence boundary revealed that the destabilizing activity of the MATalpha1 IE is observed when the terminal codon of the element's rare-codon interval is translated. This region of stability transition corresponds precisely to a MATalpha1 IE sequence previously shown to be complementary to 18S rRNA. Deletion of three nucleotides 3' to this sequence shifted the stability boundary one codon 5' to its wild-type location. Conversely, constructs containing an additional three nucleotides at this same location shifted the transition downstream by an equivalent sequence distance. Our results suggest a model in which the triggering of MATalpha1 mRNA destabilization results from establishment of an interaction between translating ribosomes and a downstream sequence element. Furthermore, our data provide direct molecular evidence for a relationship between mRNA turnover and mRNA translation.

MeSH Terms
Alleles Base Sequence Codon Fungal Proteins/metabolism Genes, Fungal Mating Factor Molecular Sequence Data Mutagenesis, Site-Directed Oligodeoxyribonucleotides Peptide Biosynthesis Peptide Chain Termination, Translational Peptides/metabolism Pheromones Plasmids Protein Biosynthesis RNA Helicases RNA, Messenger/metabolism Recombinant Proteins/metabolism Saccharomyces cerevisiae/genetics,metabolism Saccharomyces cerevisiae Proteins Transcription, Genetic
Chemicals
Codon Fungal Proteins Oligodeoxyribonucleotides Peptides Pheromones RNA, Messenger Recombinant Proteins Saccharomyces cerevisiae Proteins Mating Factor NAM7 protein, S cerevisiae RNA Helicases
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Hennigan A N
Department of Molecular Genetics and Microbiology, University of Massachusetts Medical School, Worcester, 01655-0122, USA.
Jacobson A
References (57)
57 references, click to expand
  1. Early meiotic transcripts are highly unstable in Saccharomyces cerevisiae.
    Mol Cell Biol. 1992 Sep;12(9):3948-58 PMID: 1508196
  2. Identification of pre-mRNA polyadenylation sites in Saccharomyces cerevisiae.
    Mol Cell Biol. 1992 Sep;12(9):4215-29 PMID: 1508215
  3. Mutations affecting stability and deadenylation of the yeast MFA2 transcript.
    Genes Dev. 1992 Nov;6(11):2100-11 PMID: 1427074
  4. Analysis of chimeric mRNAs derived from the STE3 mRNA identifies multiple regions within yeast mRNAs that modulate mRNA decay.
    Nucleic Acids Res. 1992 Oct 25;20(20):5365-73 PMID: 1437553
  5. A mutation in the tRNA nucleotidyltransferase gene promotes stabilization of mRNAs in Saccharomyces cerevisiae.
    Mol Cell Biol. 1992 Dec;12(12):5778-84 PMID: 1448105
  6. mRNA stability: in trans-it.
    Curr Opin Cell Biol. 1992 Dec;4(6):979-83 PMID: 1485969
  7. A turnover pathway for both stable and unstable mRNAs in yeast: evidence for a requirement for deadenylation.
    Genes Dev. 1993 Aug;7(8):1632-43 PMID: 8393418
  8. Stabilization and ribosome association of unspliced pre-mRNAs in a yeast upf1- mutant.
    Proc Natl Acad Sci U S A. 1993 Aug 1;90(15):7034-8 PMID: 8346213
  9. A small segment of the MAT alpha 1 transcript promotes mRNA decay in Saccharomyces cerevisiae: a stimulatory role for rare codons.
    Mol Cell Biol. 1993 Sep;13(9):5141-8 PMID: 8355674
  10. mRNA destabilization triggered by premature translational termination depends on at least three cis-acting sequence elements and one trans-acting factor.
    Genes Dev. 1993 Sep;7(9):1737-54 PMID: 8370523
  11. The 5' untranslated region of the PPR1 regulatory gene dictates rapid mRNA decay in yeast.
    Gene. 1993 Sep 6;131(1):43-51 PMID: 8370540
  12. Signals that produce 3' termini in CYC1 mRNA of the yeast Saccharomyces cerevisiae.
    Mol Cell Biol. 1993 Dec;13(12):7836-49 PMID: 8246998
  13. Saturation mutagenesis of a polyadenylation signal reveals a hexanucleotide element essential for mRNA 3' end formation in Saccharomyces cerevisiae.
    Proc Natl Acad Sci U S A. 1994 Jan 4;91(1):257-61 PMID: 8278376
  14. Differential effects of translational inhibition in cis and in trans on the decay of the unstable yeast MFA2 mRNA.
    J Biol Chem. 1994 Apr 1;269(13):9687-92 PMID: 8144558
  15. Nonsense-mediated mRNA decay in yeast.
    Prog Nucleic Acid Res Mol Biol. 1994;47:271-98 PMID: 8016322
  16. Deadenylation of the unstable mRNA encoded by the yeast MFA2 gene leads to decapping followed by 5'-->3' digestion of the transcript.
    Genes Dev. 1994 Apr 1;8(7):855-66 PMID: 7926773
  17. Mechanisms of mRNA degradation in eukaryotes.
    Trends Biochem Sci. 1994 Aug;19(8):336-40 PMID: 7940679
  18. Purification of a human polyribosome-associated 3' to 5' exoribonuclease.
    J Biol Chem. 1994 Dec 16;269(50):31814-21 PMID: 7989354
  19. Initiation of translation can occur only in a restricted region of the CYC1 mRNA of Saccharomyces cerevisiae.
    Mol Cell Biol. 1995 Feb;15(2):1021-33 PMID: 7823918
  20. Characterization of cis-acting sequences and decay intermediates involved in nonsense-mediated mRNA turnover.
    Mol Cell Biol. 1995 Feb;15(2):809-23 PMID: 7823948
  21. Identification and characterization of genes that are required for the accelerated degradation of mRNAs containing a premature translational termination codon.
    Genes Dev. 1995 Feb 15;9(4):423-36 PMID: 7883167
  22. Identification of a novel component of the nonsense-mediated mRNA decay pathway by use of an interacting protein screen.
    Genes Dev. 1995 Feb 15;9(4):437-54 PMID: 7883168
  23. Identification and characterization of a sequence motif involved in nonsense-mediated mRNA decay.
    Mol Cell Biol. 1995 Apr;15(4):2231-44 PMID: 7891717
  24. A complex unidirectional signal element mediates GCN4 mRNA 3' end formation in Saccharomyces cerevisiae.
    Mol Cell Biol. 1995 May;15(5):2466-73 PMID: 7739531
  25. The majority of yeast UPF1 co-localizes with polyribosomes in the cytoplasm.
    Mol Biol Cell. 1995 May;6(5):611-25 PMID: 7545033
  26. Identification of the cis-elements mediating the autogenous control of ribosomal protein L2 mRNA stability in yeast.
    EMBO J. 1995 Aug 15;14(16):4022-30 PMID: 7664741
  27. mRNA stability in mammalian cells.
    Microbiol Rev. 1995 Sep;59(3):423-50 PMID: 7565413
  28. Identification of an additional gene required for eukaryotic nonsense mRNA turnover.
    Proc Natl Acad Sci U S A. 1995 Oct 24;92(22):10354-8 PMID: 7479783
  29. Glucose-dependent turnover of the mRNAs encoding succinate dehydrogenase peptides in Saccharomyces cerevisiae: sequence elements in the 5' untranslated region of the Ip mRNA play a dominant role.
    Mol Biol Cell. 1995 Sep;6(9):1125-43 PMID: 8534911
  30. AU-rich elements: characterization and importance in mRNA degradation.
    Trends Biochem Sci. 1995 Nov;20(11):465-70 PMID: 8578590
  31. Interrelationships of the pathways of mRNA decay and translation in eukaryotic cells.
    Annu Rev Biochem. 1996;65:693-739 PMID: 8811193
  32. DNA sequencing with chain-terminating inhibitors.
    Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463-7 PMID: 271968
  33. Interference of nonsense mutations with eukaryotic messenger RNA stability.
    Proc Natl Acad Sci U S A. 1979 Oct;76(10):5134-7 PMID: 388431
  34. DNA sequence required for efficient transcription termination in yeast.
    Cell. 1982 Mar;28(3):563-73 PMID: 6280875
  35. New M13 vectors for cloning.
    Methods Enzymol. 1983;101:20-78 PMID: 6310323
  36. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity.
    Anal Biochem. 1983 Jul 1;132(1):6-13 PMID: 6312838
  37. Sequences responsible for transcription termination on a gene segment in Saccharomyces cerevisiae.
    Mol Cell Biol. 1984 Aug;4(8):1515-20 PMID: 6436686
  38. A mutation allowing an mRNA secondary structure diminishes translation of Saccharomyces cerevisiae iso-1-cytochrome c.
    Mol Cell Biol. 1985 Aug;5(8):1839-46 PMID: 3018530
  39. Purification and fractionation of poly(A)+ RNA.
    Methods Enzymol. 1987;152:254-61 PMID: 2443797
  40. Rapid and efficient site-specific mutagenesis without phenotypic selection.
    Methods Enzymol. 1987;154:367-82 PMID: 3323813
  41. Ribosome pausing and stacking during translation of a eukaryotic mRNA.
    EMBO J. 1988 Nov;7(11):3559-69 PMID: 2850168
  42. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae.
    Genetics. 1989 May;122(1):19-27 PMID: 2659436
  43. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier.
    Curr Genet. 1989 Dec;16(5-6):339-46 PMID: 2692852
  44. Translation and a 42-nucleotide segment within the coding region of the mRNA encoded by the MAT alpha 1 gene are involved in promoting rapid mRNA decay in yeast.
    Proc Natl Acad Sci U S A. 1990 Apr;87(7):2780-4 PMID: 2181450
  45. Nuclease activity associated with mammalian mRNA in its native state: possible basis for selectivity in mRNA decay.
    Mol Cell Biol. 1990 May;10(5):2060-9 PMID: 2325645
  46. Identification and comparison of stable and unstable mRNAs in Saccharomyces cerevisiae.
    Mol Cell Biol. 1990 May;10(5):2269-84 PMID: 2183028
  47. Signal sequence for generation of mRNA 3' end in the Saccharomyces cerevisiae GAL7 gene.
    EMBO J. 1990 Nov;9(11):3691-7 PMID: 2209557
  48. Yeast Saccharomyces cerevisiae selectable markers in pUC18 polylinkers.
    Yeast. 1990 Sep-Oct;6(5):363-6 PMID: 2220072
  49. Translation of c-myc mRNA is required for its post-transcriptional regulation during myogenesis.
    J Biol Chem. 1990 Nov 5;265(31):19015-21 PMID: 2229059
  50. 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):221-31 PMID: 1899842
  51. Distinct cis-acting signals enhance 3' endpoint formation of CYC1 mRNA in the yeast Saccharomyces cerevisiae.
    EMBO J. 1991 Mar;10(3):563-71 PMID: 1848175
  52. Different classes of polyadenylation sites in the yeast Saccharomyces cerevisiae.
    Mol Cell Biol. 1991 Jun;11(6):3060-9 PMID: 2038317
  53. The product of the yeast UPF1 gene is required for rapid turnover of mRNAs containing a premature translational termination codon.
    Genes Dev. 1991 Dec;5(12A):2303-14 PMID: 1748286
  54. Regulation of mRNA turnover in eukaryotic cells.
    Crit Rev Eukaryot Gene Expr. 1991;1(2):99-126 PMID: 1802106
  55. Gene products that promote mRNA turnover in Saccharomyces cerevisiae.
    Mol Cell Biol. 1992 May;12(5):2165-77 PMID: 1569946
  56. Different sequence elements are required for function of the cauliflower mosaic virus polyadenylation site in Saccharomyces cerevisiae compared with in plants.
    Mol Cell Biol. 1992 May;12(5):2322-30 PMID: 1373813
  57. A coding region segment is necessary, but not sufficient for rapid decay of the HIS3 mRNA in yeast.
    Gene. 1992 May 1;114(1):35-41 PMID: 1587483
Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.16.7.3833
Published
1996-07-00
Pages
3833-43
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC231380
Subset
IM
Grants
NIGMS NIH HHS · R01 GM027757 · United States
NIGMS NIH HHS · GM27757 · United States
External Links
PubMed source DOI: 10.1128/MCB.16.7.3833 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2