1887

Abstract

The capsids of ssRNA phages comprise a single copy of an ~45 kDa maturation protein that serves to recognize the conjugative pilus as receptor, to protect the ends of the viral RNA and also to escort the genomic RNA into the host cytoplasm. In the , represented by the canonical phage Qβ, the maturation protein A also causes lysis. This is achieved by inhibiting the activity of MurA, which catalyses the first committed step of murein biosynthesis. Previously, it was shown that Qβ virions, with a single copy of A, inhibit MurA activity. This led to a model for lysis timing in which, during phage infection, A is not active as a MurA inhibitor until assembled into virion particles, thus preventing premature lysis before a sufficient yield of viable progeny has accumulated. Here we report that MurA inactivates purified Qβ particles, casting doubt on the notion that A must assemble into particles prior to MurA inhibition. Furthermore, quantification of A protein induced from a plasmid indicated that lysis is entrained when the amount of the lysis protein is approximately equimolar to that of cellular MurA. Qβ mutants, isolated as suppressors that overcome a mutation that reduces the affinity of MurA for A, were shown to be missense mutations in that increase the translation of the maturation protein. Because of the increased production of A, the mutants have an attenuated infection cycle and reduced burst size, indicating that a delicate balance between assembled and unassembled A levels regulates lysis timing.

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2013-03-01
2020-10-20
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References

  1. Beekwilder J., Nieuwenhuizen R., Poot R., van Duin J.. ( 1996;). Secondary structure model for the first three domains of Qβ RNA. Control of A-protein synthesis. J Mol Biol256:8–19[PubMed][CrossRef]
    [Google Scholar]
  2. Bernhardt T. G., Roof W. D., Young R.. ( 2000;). Genetic evidence that the bacteriophage ϕ X174 lysis protein inhibits cell wall synthesis. Proc Natl Acad Sci U S A97:4297–4302 [CrossRef][PubMed]
    [Google Scholar]
  3. Bernhardt T. G., Wang I. N., Struck D. K., Young R.. ( 2001;). A protein antibiotic in the phage Qβ virion: diversity in lysis targets. Science292:2326–2329 [CrossRef][PubMed]
    [Google Scholar]
  4. Bernhardt T. G., Wang I. N., Struck D. K., Young R.. ( 2002;). Breaking free: “protein antibiotics” and phage lysis. Res Microbiol153:493–501 [CrossRef][PubMed]
    [Google Scholar]
  5. Blumenthal T., Carmichael G. G.. ( 1979;). RNA replication: function and structure of Qβ-replicase. Annu Rev Biochem48:525–548 [CrossRef][PubMed]
    [Google Scholar]
  6. Boedtker H., Gesteland R. F.. ( 1975;). Physical properties. RNA Phages Zinder N. D.. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  7. Brown E. D., Vivas E. I., Walsh C. T., Kolter R.. ( 1995;). MurA (MurZ), the enzyme that catalyzes the first committed step in peptidoglycan biosynthesis, is essential in Escherichia coli . J Bacteriol177:4194–4197[PubMed]
    [Google Scholar]
  8. Domingo E., Holland J. J.. ( 1997;). RNA virus mutations and fitness for survival. Annu Rev Microbiol51:151–178 [CrossRef][PubMed]
    [Google Scholar]
  9. Domingo E., Sabo D., Taniguchi T., Weissmann C.. ( 1978;). Nucleotide sequence heterogeneity of an RNA phage population. Cell13:735–744 [CrossRef][PubMed]
    [Google Scholar]
  10. Drake J. W.. ( 1993;). Rates of spontaneous mutation among RNA viruses. Proc Natl Acad Sci U S A90:4171–4175 [CrossRef][PubMed]
    [Google Scholar]
  11. Dykeman E. C., Grayson N. E., Toropova K., Ranson N. A., Stockley P. G., Twarock R.. ( 2011;). Simple rules for efficient assembly predict the layout of a packaged viral RNA. J Mol Biol408:399–407 [CrossRef][PubMed]
    [Google Scholar]
  12. Groeneveld H., Thimon K., van Duin J.. ( 1995;). Translational control of maturation-protein synthesis in phage MS2: a role for the kinetics of RNA folding?. RNA1:79–88[PubMed]
    [Google Scholar]
  13. Gründling A., Bläsi U., Young R.. ( 2000;). Biochemical and genetic evidence for three transmembrane domains in the class I holin, λ S. J Biol Chem275:769–776 [CrossRef][PubMed]
    [Google Scholar]
  14. Hatfull G. F.. ( 2001;). Microbiology. The great escape. Science292:2263–2264 [CrossRef][PubMed]
    [Google Scholar]
  15. Ishihama Y., Schmidt T., Rappsilber J., Mann M., Hartl F. U., Kerner M. J., Frishman D.. ( 2008;). Protein abundance profiling of the Escherichia coli cytosol. BMC Genomics9:102[PubMed][CrossRef]
    [Google Scholar]
  16. Jensen P. R., Westerhoff H. V., Michelsen O.. ( 1993;). The use of lac-type promoters in control analysis. Eur J Biochem211:181–191[PubMed][CrossRef]
    [Google Scholar]
  17. Kozak M., Nathans D.. ( 1971;). Fate of maturation protein during infection by coliphage MS2. Nat New Biol234:209–211[PubMed][CrossRef]
    [Google Scholar]
  18. Palmer I., Wingfield P. T.. ( 2001;). Preparation and extraction of insoluble (inclusion-body) proteins from Escherichia coli . Current Protocols in Protein Science70:6.3.1–6.3.20 [CrossRef]
    [Google Scholar]
  19. Reed C. A., Langlais C., Kuznetsov V., Young R.. ( 2012;). Inhibitory mechanism of the Qβ lysis protein A2 . Mol Microbiol86:836–844[CrossRef]
    [Google Scholar]
  20. Riley-Lovingshimer M. R., Reinhart G. D.. ( 2001;). Equilibrium binding studies of a tryptophan-shifted mutant of phosphofructokinase from Bacillus stearothermophilus . Biochemistry40:3002–3008[PubMed][CrossRef]
    [Google Scholar]
  21. Robertson H. D., Lodish H. F.. ( 1970;). Messenger characteristics of nascent bacteriophage RNA. Proc Natl Acad Sci U S A67:710–716 [CrossRef][PubMed]
    [Google Scholar]
  22. Sambrook J., Fritsch E. F., Maniatis T., Sambrook J., Fritsch E. F., Maniatis T.. ( 1989;). Molecular Cloning: a Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  23. Shiba T., Suzuki Y.. ( 1981;). Localization of A protein in the RNA-A protein complex of RNA phage MS2. Biochim Biophys Acta654:249–255 [CrossRef][PubMed]
    [Google Scholar]
  24. Siegele D. A., Hu J. C.. ( 1997;). Gene expression from plasmids containing the araBAD promoter at subsaturating inducer concentrations represents mixed populations. Proc Natl Acad Sci U S A94:8168–8172[PubMed][CrossRef]
    [Google Scholar]
  25. Staples D. H., Hindley J., Billeter M. A., Weissmann C.. ( 1971;). Localization of Q-β maturation cistron ribosome binding site. Nat New Biol234:202–204[PubMed][CrossRef]
    [Google Scholar]
  26. Strauss J. H. Jr, Sinsheimer R. L.. ( 1963;). Purification and properties of bacteriophage MS2 and of its ribonucleic acid. J Mol Biol7:43–54 [CrossRef][PubMed]
    [Google Scholar]
  27. Tsukada K., Okazaki M., Kita H., Inokuchi Y., Urabe I., Yomo T.. ( 2009;). Quantitative analysis of the bacteriophage Qβ infection cycle. Biochim Biophys Acta1790:65–70 [CrossRef]
    [Google Scholar]
  28. van Duin J., Tsareva N.. ( 2006;). Single-stranded RNA phages. The Bacteriophages175–196 Calendar R.. Oxford: Oxford University Press;
    [Google Scholar]
  29. Weber K., Konigsberg W., Zinder N. D.. ( 1975;). Proteins of the RNA phages. RNA Phages51–84 Zinder N. D.. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  30. Winter R. B., Gold L.. ( 1983;). Overproduction of bacteriophage Q β maturation (A2) protein leads to cell lysis. Cell33:877–885 [CrossRef][PubMed]
    [Google Scholar]
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