Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 May;178(10):2926–2933. doi: 10.1128/jb.178.10.2926-2933.1996

Overexpression of an mRNA dependent on rare codons inhibits protein synthesis and cell growth.

K Zahn 1
PMCID: PMC178030  PMID: 8631683

Abstract

lambda's int gene contains an unusually high frequency of the rare arginine codons AGA and AGG, as well as dual rare Arg codons at three positions. Related work has demonstrated that Int protein expression depends on the rare AGA tRNA. Strong transcription of the int mRNA with a highly efficient ribosome-binding site leads to inhibition of Int protein synthesis, alteration of the overall pattern of cellular protein synthesis, and cell death. Synthesis or stability of int and ampicillin resistance mRNAs is not affected, although a portion of the untranslated int mRNA appears to be modified in a site-specific fashion. These phenotypes are not due to a toxic effect of the int gene product and can be largely reversed by supplementation of the AGA tRNA in cells which bear plasmids expressing the T4 AGA tRNA gene. This indicates that depletion of the rare Arg tRNA due to ribosome stalling at multiple AGA and AGG codons on the overexpressed int mRNA underlies all of these phenomena. It is hypothesized that int mRNA's effects on protein synthesis and cell viability relate to phenomena involved in lambda phage induction and excision.

Full Text

The Full Text of this article is available as a PDF (540.2 KB).

Selected References

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

  1. Aota S., Gojobori T., Ishibashi F., Maruyama T., Ikemura T. Codon usage tabulated from the GenBank Genetic Sequence Data. Nucleic Acids Res. 1988;16 (Suppl):r315–r402. doi: 10.1093/nar/16.suppl.r315. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bonekamp F., Jensen K. F. The AGG codon is translated slowly in E. coli even at very low expression levels. Nucleic Acids Res. 1988 Apr 11;16(7):3013–3024. doi: 10.1093/nar/16.7.3013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brinkmann U., Mattes R. E., Buckel P. High-level expression of recombinant genes in Escherichia coli is dependent on the availability of the dnaY gene product. Gene. 1989 Dec 21;85(1):109–114. doi: 10.1016/0378-1119(89)90470-8. [DOI] [PubMed] [Google Scholar]
  4. Chang A. C., Cohen S. N. Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol. 1978 Jun;134(3):1141–1156. doi: 10.1128/jb.134.3.1141-1156.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chen G. F., Inouye M. Suppression of the negative effect of minor arginine codons on gene expression; preferential usage of minor codons within the first 25 codons of the Escherichia coli genes. Nucleic Acids Res. 1990 Mar 25;18(6):1465–1473. doi: 10.1093/nar/18.6.1465. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chen G. T., Inouye M. Role of the AGA/AGG codons, the rarest codons in global gene expression in Escherichia coli. Genes Dev. 1994 Nov 1;8(21):2641–2652. doi: 10.1101/gad.8.21.2641. [DOI] [PubMed] [Google Scholar]
  7. Chen K. S., Peters T. C., Walker J. R. A minor arginine tRNA mutant limits translation preferentially of a protein dependent on the cognate codon. J Bacteriol. 1990 May;172(5):2504–2510. doi: 10.1128/jb.172.5.2504-2510.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cohen S. N., Chang A. C. Genetic expression in bacteriophage lambda. 3. Inhibition of Escherichia coli nucleic acid and protein synthesis during lambda development. J Mol Biol. 1970 May 14;49(3):557–575. doi: 10.1016/0022-2836(70)90281-0. [DOI] [PubMed] [Google Scholar]
  9. Daniels D. L., Sanger F., Coulson A. R. Features of bacteriophage lambda: analysis of the complete nucleotide sequence. Cold Spring Harb Symp Quant Biol. 1983;47(Pt 2):1009–1024. doi: 10.1101/sqb.1983.047.01.115. [DOI] [PubMed] [Google Scholar]
  10. Dong H., Nilsson L., Kurland C. G. Gratuitous overexpression of genes in Escherichia coli leads to growth inhibition and ribosome destruction. J Bacteriol. 1995 Mar;177(6):1497–1504. doi: 10.1128/jb.177.6.1497-1504.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Emilsson V., Näslund A. K., Kurland C. G. Growth-rate-dependent accumulation of twelve tRNA species in Escherichia coli. J Mol Biol. 1993 Mar 20;230(2):483–491. doi: 10.1006/jmbi.1993.1165. [DOI] [PubMed] [Google Scholar]
  12. Garcia G. M., Mar P. K., Mullin D. A., Walker J. R., Prather N. E. The E. coli dnaY gene encodes an arginine transfer RNA. Cell. 1986 May 9;45(3):453–459. doi: 10.1016/0092-8674(86)90331-4. [DOI] [PubMed] [Google Scholar]
  13. Gough J. A., Murray N. E. Sequence diversity among related genes for recognition of specific targets in DNA molecules. J Mol Biol. 1983 May 5;166(1):1–19. doi: 10.1016/s0022-2836(83)80047-3. [DOI] [PubMed] [Google Scholar]
  14. Guarneros G., Montañez C., Hernandez T., Court D. Posttranscriptional control of bacteriophage lambda gene expression from a site distal to the gene. Proc Natl Acad Sci U S A. 1982 Jan;79(2):238–242. doi: 10.1073/pnas.79.2.238. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Guzman P., Rivera Chavira B. E., Court D. L., Gottesman M. E., Guarneros G. Transcription of a bacteriophage lambda DNA site blocks growth of Escherichia coli. J Bacteriol. 1990 Feb;172(2):1030–1034. doi: 10.1128/jb.172.2.1030-1034.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Henson J. M., Chu H., Irwin C. A., Walker J. R. Isolation and characterization of dnaX and dnaY temperature-sensitive mutants of Escherichia coli. Genetics. 1979 Aug;92(4):1041–1059. doi: 10.1093/genetics/92.4.1041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Ikemura T. Correlation between the abundance of Escherichia coli transfer RNAs and the occurrence of the respective codons in its protein genes: a proposal for a synonymous codon choice that is optimal for the E. coli translational system. J Mol Biol. 1981 Sep 25;151(3):389–409. doi: 10.1016/0022-2836(81)90003-6. [DOI] [PubMed] [Google Scholar]
  18. Kikuchi Y., Nash H. A. Nicking-closing activity associated with bacteriophage lambda int gene product. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3760–3764. doi: 10.1073/pnas.76.8.3760. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lorsch J. R., Bartel D. P., Szostak J. W. Reverse transcriptase reads through a 2'-5'linkage and a 2'-thiophosphate in a template. Nucleic Acids Res. 1995 Aug 11;23(15):2811–2814. doi: 10.1093/nar/23.15.2811. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  21. Mazzara G. P., Plunkett G., 3rd, McClain W. H. DNA sequence of the transfer RNA region of bacteriophage T4: implications for transfer RNA synthesis. Proc Natl Acad Sci U S A. 1981 Feb;78(2):889–892. doi: 10.1073/pnas.78.2.889. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Misra R., Reeves P. Intermediates in the synthesis of TolC protein include an incomplete peptide stalled at a rare Arg codon. Eur J Biochem. 1985 Oct 1;152(1):151–155. doi: 10.1111/j.1432-1033.1985.tb09175.x. [DOI] [PubMed] [Google Scholar]
  23. Montañez C., Bueno J., Schmeissner U., Court D. L., Guarneros G. Mutations of bacteriophage lambda that define independent but overlapping RNA processing and transcription termination sites. J Mol Biol. 1986 Sep 5;191(1):29–37. doi: 10.1016/0022-2836(86)90420-1. [DOI] [PubMed] [Google Scholar]
  24. Nunes-Düby S. E., Matsumoto L., Landy A. Site-specific recombination intermediates trapped with suicide substrates. Cell. 1987 Aug 28;50(5):779–788. doi: 10.1016/0092-8674(87)90336-9. [DOI] [PubMed] [Google Scholar]
  25. Oakley B. R., Kirsch D. R., Morris N. R. A simplified ultrasensitive silver stain for detecting proteins in polyacrylamide gels. Anal Biochem. 1980 Jul 1;105(2):361–363. doi: 10.1016/0003-2697(80)90470-4. [DOI] [PubMed] [Google Scholar]
  26. Olins P. O., Rangwala S. H. A novel sequence element derived from bacteriophage T7 mRNA acts as an enhancer of translation of the lacZ gene in Escherichia coli. J Biol Chem. 1989 Oct 15;264(29):16973–16976. [PubMed] [Google Scholar]
  27. Oppenheim A. B., Gottesman S., Gottesman M. Regulation of bacteriophage lambda int gene expression. J Mol Biol. 1982 Jul 5;158(3):327–346. doi: 10.1016/0022-2836(82)90201-7. [DOI] [PubMed] [Google Scholar]
  28. Pargellis C. A., Nunes-Düby S. E., de Vargas L. M., Landy A. Suicide recombination substrates yield covalent lambda integrase-DNA complexes and lead to identification of the active site tyrosine. J Biol Chem. 1988 Jun 5;263(16):7678–7685. [PubMed] [Google Scholar]
  29. Plunkett G., 3rd, Echols H. Retroregulation of the bacteriophage lambda int gene: limited secondary degradation of the RNase III-processed transcript. J Bacteriol. 1989 Jan;171(1):588–592. doi: 10.1128/jb.171.1.588-592.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Pérez-Morga D., Guarneros G. A short DNA sequence from lambda phage inhibits protein synthesis in Escherichia coli rap. J Mol Biol. 1990 Nov 20;216(2):243–250. doi: 10.1016/s0022-2836(05)80316-x. [DOI] [PubMed] [Google Scholar]
  31. Reyes O., Gottesman M., Adhya S. Suppression of polarity of insertion mutations in the gal operon and N mutations in bacteriophage lambda. J Bacteriol. 1976 Jun;126(3):1108–1112. doi: 10.1128/jb.126.3.1108-1112.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Saxena P., Walker J. R. Expression of argU, the Escherichia coli gene coding for a rare arginine tRNA. J Bacteriol. 1992 Mar;174(6):1956–1964. doi: 10.1128/jb.174.6.1956-1964.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Simatake H., Rosenberg M. Purified lambda regulatory protein cII positively activates promoters for lysogenic development. Nature. 1981 Jul 9;292(5819):128–132. doi: 10.1038/292128a0. [DOI] [PubMed] [Google Scholar]
  34. Spanjaard R. A., van Duin J. Translation of the sequence AGG-AGG yields 50% ribosomal frameshift. Proc Natl Acad Sci U S A. 1988 Nov;85(21):7967–7971. doi: 10.1073/pnas.85.21.7967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Sørensen M. A., Kurland C. G., Pedersen S. Codon usage determines translation rate in Escherichia coli. J Mol Biol. 1989 May 20;207(2):365–377. doi: 10.1016/0022-2836(89)90260-x. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES