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. 1995 Jul;177(13):3695–3703. doi: 10.1128/jb.177.13.3695-3703.1995

An essential role for the Escherichia coli DnaK protein in starvation-induced thermotolerance, H2O2 resistance, and reductive division.

D Rockabrand 1, T Arthur 1, G Korinek 1, K Livers 1, P Blum 1
PMCID: PMC177085  PMID: 7601833

Abstract

During a 3-day period, glucose starvation of wild-type Escherichia coli produced thermotolerant, H2O2-resistant, small cells with a round morphology. These cells contained elevated levels of the DnaK protein, adjusted either for total protein or on a per-cell basis. Immunoprecipitation of [35S]methionine-labeled protein produced by such starving cells demonstrated that DnaK underwent continuous synthesis but at decreasing rates throughout this time. Glucose resupplementation of starving cells resulted in rapid loss of thermotolerance, H2O2 resistance, and the elevated DnaK levels. A dnaK deletion mutant, but not an otherwise isogenic wild-type strain, failed to develop starvation-induced thermotolerance or H2O2 resistance. The filamentous phenotype associated with DnaK deficiency was suppressed by cultivation in a defined glucose medium. When starved for glucose, the nonfilamentous and rod-shaped dnaK mutant strain failed to convert into the small spherical form typical of starving wild-type cells. The dnaK mutant retained the ability to develop adaptive H2O2 resistance during growth but not adaptive resistance to heat. Complementation of DnaK deficiency by using Ptac-regulated dnaK+ and dnaK+J+ expression plasmids confirmed a specific role for the DnaK molecular chaperone in these starvation-induced phenotypes.

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Selected References

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  1. Aldea M., Garrido T., Hernández-Chico C., Vicente M., Kushner S. R. Induction of a growth-phase-dependent promoter triggers transcription of bolA, an Escherichia coli morphogene. EMBO J. 1989 Dec 1;8(12):3923–3931. doi: 10.1002/j.1460-2075.1989.tb08573.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Almirón M., Link A. J., Furlong D., Kolter R. A novel DNA-binding protein with regulatory and protective roles in starved Escherichia coli. Genes Dev. 1992 Dec;6(12B):2646–2654. doi: 10.1101/gad.6.12b.2646. [DOI] [PubMed] [Google Scholar]
  3. Blum P. H., Jovanovich S. B., McCann M. P., Schultz J. E., Lesley S. A., Burgess R. R., Matin A. Cloning and in vivo and in vitro regulation of cyclic AMP-dependent carbon starvation genes from Escherichia coli. J Bacteriol. 1990 Jul;172(7):3813–3820. doi: 10.1128/jb.172.7.3813-3820.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Blum P., Ory J., Bauernfeind J., Krska J. Physiological consequences of DnaK and DnaJ overproduction in Escherichia coli. J Bacteriol. 1992 Nov;174(22):7436–7444. doi: 10.1128/jb.174.22.7436-7444.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bukau B., Walker G. C. Cellular defects caused by deletion of the Escherichia coli dnaK gene indicate roles for heat shock protein in normal metabolism. J Bacteriol. 1989 May;171(5):2337–2346. doi: 10.1128/jb.171.5.2337-2346.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bukau B., Walker G. C. Delta dnaK52 mutants of Escherichia coli have defects in chromosome segregation and plasmid maintenance at normal growth temperatures. J Bacteriol. 1989 Nov;171(11):6030–6038. doi: 10.1128/jb.171.11.6030-6038.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bukau B., Walker G. C. Mutations altering heat shock specific subunit of RNA polymerase suppress major cellular defects of E. coli mutants lacking the DnaK chaperone. EMBO J. 1990 Dec;9(12):4027–4036. doi: 10.1002/j.1460-2075.1990.tb07624.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Burkholder W. F., Panagiotidis C. A., Silverstein S. J., Cegielska A., Gottesman M. E., Gaitanaris G. A. Isolation and characterization of an Escherichia coli DnaK mutant with impaired ATPase activity. J Mol Biol. 1994 Sep 30;242(4):364–377. doi: 10.1006/jmbi.1994.1587. [DOI] [PubMed] [Google Scholar]
  9. Dean D. O., James R. Identification of a gene, closely linked to dnaK, which is required for high-temperature growth of Escherichia coli. J Gen Microbiol. 1991 Jun;137(6):1271–1277. doi: 10.1099/00221287-137-6-1271. [DOI] [PubMed] [Google Scholar]
  10. Delaney J. M. Requirement of the Escherichia coli dnaK gene for thermotolerance and protection against H2O2. J Gen Microbiol. 1990 Oct;136(10):2113–2118. doi: 10.1099/00221287-136-10-2113. [DOI] [PubMed] [Google Scholar]
  11. Demple B. Regulation of bacterial oxidative stress genes. Annu Rev Genet. 1991;25:315–337. doi: 10.1146/annurev.ge.25.120191.001531. [DOI] [PubMed] [Google Scholar]
  12. Elliker P. R., Frazier W. C. Influence of Time and Temperature of Incubation on Heat Resistance of Escherichia coli. J Bacteriol. 1938 Jul;36(1):83–98. doi: 10.1128/jb.36.1.83-98.1938. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Erickson J. W., Gross C. A. Identification of the sigma E subunit of Escherichia coli RNA polymerase: a second alternate sigma factor involved in high-temperature gene expression. Genes Dev. 1989 Sep;3(9):1462–1471. doi: 10.1101/gad.3.9.1462. [DOI] [PubMed] [Google Scholar]
  14. Georgopoulos C. The emergence of the chaperone machines. Trends Biochem Sci. 1992 Aug;17(8):295–299. doi: 10.1016/0968-0004(92)90439-g. [DOI] [PubMed] [Google Scholar]
  15. Giraldo-Suárez R., Fernández-Tresguerres E., Díaz-Orejas R., Malki A., Kohiyama M. The heat-shock DnaK protein is required for plasmid R1 replication and it is dispensable for plasmid ColE1 replication. Nucleic Acids Res. 1993 Nov 25;21(23):5495–5499. doi: 10.1093/nar/21.23.5495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Groat R. G., Schultz J. E., Zychlinsky E., Bockman A., Matin A. Starvation proteins in Escherichia coli: kinetics of synthesis and role in starvation survival. J Bacteriol. 1986 Nov;168(2):486–493. doi: 10.1128/jb.168.2.486-493.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Grossman A. D., Taylor W. E., Burton Z. F., Burgess R. R., Gross C. A. Stringent response in Escherichia coli induces expression of heat shock proteins. J Mol Biol. 1985 Nov 20;186(2):357–365. doi: 10.1016/0022-2836(85)90110-x. [DOI] [PubMed] [Google Scholar]
  18. Hengge-Aronis R., Klein W., Lange R., Rimmele M., Boos W. Trehalose synthesis genes are controlled by the putative sigma factor encoded by rpoS and are involved in stationary-phase thermotolerance in Escherichia coli. J Bacteriol. 1991 Dec;173(24):7918–7924. doi: 10.1128/jb.173.24.7918-7924.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. James R., Dean D. O., Debbage J. Five open reading frames upstream of the dnaK gene of E. coli. DNA Seq. 1993;3(5):327–332. doi: 10.3109/10425179309020832. [DOI] [PubMed] [Google Scholar]
  20. Jenkins D. E., Auger E. A., Matin A. Role of RpoH, a heat shock regulator protein, in Escherichia coli carbon starvation protein synthesis and survival. J Bacteriol. 1991 Mar;173(6):1992–1996. doi: 10.1128/jb.173.6.1992-1996.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jenkins D. E., Schultz J. E., Matin A. Starvation-induced cross protection against heat or H2O2 challenge in Escherichia coli. J Bacteriol. 1988 Sep;170(9):3910–3914. doi: 10.1128/jb.170.9.3910-3914.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kawamukai M., Matsuda H., Fujii W., Utsumi R., Komano T. Nucleotide sequences of fic and fic-1 genes involved in cell filamentation induced by cyclic AMP in Escherichia coli. J Bacteriol. 1989 Aug;171(8):4525–4529. doi: 10.1128/jb.171.8.4525-4529.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kawula T. H., Lelivelt M. J. Mutations in a gene encoding a new Hsp70 suppress rapid DNA inversion and bgl activation, but not proU derepression, in hns-1 mutant Escherichia coli. J Bacteriol. 1994 Feb;176(3):610–619. doi: 10.1128/jb.176.3.610-619.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kolter R., Siegele D. A., Tormo A. The stationary phase of the bacterial life cycle. Annu Rev Microbiol. 1993;47:855–874. doi: 10.1146/annurev.mi.47.100193.004231. [DOI] [PubMed] [Google Scholar]
  25. Krska J., Elthon T., Blum P. Monoclonal antibody recognition and function of a DnaK (HSP70) epitope found in gram-negative bacteria. J Bacteriol. 1993 Oct;175(20):6433–6440. doi: 10.1128/jb.175.20.6433-6440.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kusukawa N., Yura T. Heat shock protein GroE of Escherichia coli: key protective roles against thermal stress. Genes Dev. 1988 Jul;2(7):874–882. doi: 10.1101/gad.2.7.874. [DOI] [PubMed] [Google Scholar]
  27. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  28. Lange R., Hengge-Aronis R. Growth phase-regulated expression of bolA and morphology of stationary-phase Escherichia coli cells are controlled by the novel sigma factor sigma S. J Bacteriol. 1991 Jul;173(14):4474–4481. doi: 10.1128/jb.173.14.4474-4481.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lange R., Hengge-Aronis R. Identification of a central regulator of stationary-phase gene expression in Escherichia coli. Mol Microbiol. 1991 Jan;5(1):49–59. doi: 10.1111/j.1365-2958.1991.tb01825.x. [DOI] [PubMed] [Google Scholar]
  30. Li C., Clarke S. A protein methyltransferase specific for altered aspartyl residues is important in Escherichia coli stationary-phase survival and heat-shock resistance. Proc Natl Acad Sci U S A. 1992 Oct 15;89(20):9885–9889. doi: 10.1073/pnas.89.20.9885. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Loewen P. C., Hengge-Aronis R. The role of the sigma factor sigma S (KatF) in bacterial global regulation. Annu Rev Microbiol. 1994;48:53–80. doi: 10.1146/annurev.mi.48.100194.000413. [DOI] [PubMed] [Google Scholar]
  32. Lomovskaya O. L., Kidwell J. P., Matin A. Characterization of the sigma 38-dependent expression of a core Escherichia coli starvation gene, pexB. J Bacteriol. 1994 Jul;176(13):3928–3935. doi: 10.1128/jb.176.13.3928-3935.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Matin A., Auger E. A., Blum P. H., Schultz J. E. Genetic basis of starvation survival in nondifferentiating bacteria. Annu Rev Microbiol. 1989;43:293–316. doi: 10.1146/annurev.mi.43.100189.001453. [DOI] [PubMed] [Google Scholar]
  34. McCann M. P., Kidwell J. P., Matin A. The putative sigma factor KatF has a central role in development of starvation-mediated general resistance in Escherichia coli. J Bacteriol. 1991 Jul;173(13):4188–4194. doi: 10.1128/jb.173.13.4188-4194.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. McCarty J. S., Walker G. C. DnaK mutants defective in ATPase activity are defective in negative regulation of the heat shock response: expression of mutant DnaK proteins results in filamentation. J Bacteriol. 1994 Feb;176(3):764–780. doi: 10.1128/jb.176.3.764-780.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Meury J., Kohiyama M. Role of heat shock protein DnaK in osmotic adaptation of Escherichia coli. J Bacteriol. 1991 Jul;173(14):4404–4410. doi: 10.1128/jb.173.14.4404-4410.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Missiakas D., Georgopoulos C., Raina S. The Escherichia coli heat shock gene htpY: mutational analysis, cloning, sequencing, and transcriptional regulation. J Bacteriol. 1993 May;175(9):2613–2624. doi: 10.1128/jb.175.9.2613-2624.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Morgan R. W., Christman M. F., Jacobson F. S., Storz G., Ames B. N. Hydrogen peroxide-inducible proteins in Salmonella typhimurium overlap with heat shock and other stress proteins. Proc Natl Acad Sci U S A. 1986 Nov;83(21):8059–8063. doi: 10.1073/pnas.83.21.8059. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Paek K. H., Walker G. C. Escherichia coli dnaK null mutants are inviable at high temperature. J Bacteriol. 1987 Jan;169(1):283–290. doi: 10.1128/jb.169.1.283-290.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Pedersen S., Bloch P. L., Reeh S., Neidhardt F. C. Patterns of protein synthesis in E. coli: a catalog of the amount of 140 individual proteins at different growth rates. Cell. 1978 May;14(1):179–190. doi: 10.1016/0092-8674(78)90312-4. [DOI] [PubMed] [Google Scholar]
  41. Raina S., Missiakas D., Baird L., Kumar S., Georgopoulos C. Identification and transcriptional analysis of the Escherichia coli htrE operon which is homologous to pap and related pilin operons. J Bacteriol. 1993 Aug;175(16):5009–5021. doi: 10.1128/jb.175.16.5009-5021.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Seaton B. L., Vickery L. E. A gene encoding a DnaK/hsp70 homolog in Escherichia coli. Proc Natl Acad Sci U S A. 1994 Mar 15;91(6):2066–2070. doi: 10.1073/pnas.91.6.2066. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Shand R. F., Miercke L. J., Mitra A. K., Fong S. K., Stroud R. M., Betlach M. C. Wild-type and mutant bacterioopsins D85N, D96N, and R82Q: high-level expression in Escherichia coli. Biochemistry. 1991 Mar 26;30(12):3082–3088. doi: 10.1021/bi00226a015. [DOI] [PubMed] [Google Scholar]
  44. Spence J., Cegielska A., Georgopoulos C. Role of Escherichia coli heat shock proteins DnaK and HtpG (C62.5) in response to nutritional deprivation. J Bacteriol. 1990 Dec;172(12):7157–7166. doi: 10.1128/jb.172.12.7157-7166.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. VanBogelen R. A., Acton M. A., Neidhardt F. C. Induction of the heat shock regulon does not produce thermotolerance in Escherichia coli. Genes Dev. 1987 Aug;1(6):525–531. doi: 10.1101/gad.1.6.525. [DOI] [PubMed] [Google Scholar]
  46. VanBogelen R. A., Sankar P., Clark R. L., Bogan J. A., Neidhardt F. C. The gene-protein database of Escherichia coli: edition 5. Electrophoresis. 1992 Dec;13(12):1014–1054. doi: 10.1002/elps.11501301203. [DOI] [PubMed] [Google Scholar]
  47. Yamamori T., Yura T. Genetic control of heat-shock protein synthesis and its bearing on growth and thermal resistance in Escherichia coli K-12. Proc Natl Acad Sci U S A. 1982 Feb;79(3):860–864. doi: 10.1073/pnas.79.3.860. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Yura T., Nagai H., Mori H. Regulation of the heat-shock response in bacteria. Annu Rev Microbiol. 1993;47:321–350. doi: 10.1146/annurev.mi.47.100193.001541. [DOI] [PubMed] [Google Scholar]
  49. Zhou Y. N., Kusukawa N., Erickson J. W., Gross C. A., Yura T. Isolation and characterization of Escherichia coli mutants that lack the heat shock sigma factor sigma 32. J Bacteriol. 1988 Aug;170(8):3640–3649. doi: 10.1128/jb.170.8.3640-3649.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]

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