Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1989 May;171(5):2337–2346. doi: 10.1128/jb.171.5.2337-2346.1989

Cellular defects caused by deletion of the Escherichia coli dnaK gene indicate roles for heat shock protein in normal metabolism.

B Bukau 1, G C Walker 1
PMCID: PMC209906  PMID: 2651398

Abstract

DnaK is a major heat shock protein of Escherichia coli and has been previously reported to be essential for growth at high temperatures. We systematically investigated the role of DnaK in cellular metabolism at a wide range of growth temperatures by analyzing cellular defects caused by deletion of the dnaK gene (delta dnaK52). At intermediate temperatures (30 degrees C), introduction of the delta dnaK52 allele into wild-type cells caused severe defects in cell division, slow growth, and poor viability of the cells. delta dnaK52 mutants were genetically unstable at 30 degrees C and frequently acquired secondary mutations. At high (42 degrees C) and low (11 and 16 degrees C) temperatures the delta dnaK52 allele could only be introduced into the subpopulation of wild-type cells that had duplicated the dnaK region of their chromosome. delta dnaK52 mutants isolated at 30 degrees C were cold sensitive as well as temperature sensitive for growth. Cell division defects of delta dnaK52 mutants at 30 degrees C were largely suppressed by overproduction of the FtsZ protein, which is normally required for septation during cell division; however, slow growth and poor viability at 30 degrees C and cold sensitivity and temperature sensitivity of growth were not suppressed, indicating that delta dnaK52 mutants had additional defective cellular functions besides cell division.

Full text

PDF
2337

Images in this article

2339Image
on p.2339
2339Image
on p.2339
2341Image
on p.2341
2342Image
on p.2342
2343Image
on p.2343

Selected References

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

  1. Bardwell J. C., Craig E. A. Eukaryotic Mr 83,000 heat shock protein has a homologue in Escherichia coli. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5177–5181. doi: 10.1073/pnas.84.15.5177. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bardwell J. C., Craig E. A. Major heat shock gene of Drosophila and the Escherichia coli heat-inducible dnaK gene are homologous. Proc Natl Acad Sci U S A. 1984 Feb;81(3):848–852. doi: 10.1073/pnas.81.3.848. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bochner B. R., Zylicz M., Georgopoulos C. Escherichia coli DnaK protein possesses a 5'-nucleotidase activity that is inhibited by AppppA. J Bacteriol. 1986 Nov;168(2):931–935. doi: 10.1128/jb.168.2.931-935.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Casadaban M. J. Transposition and fusion of the lac genes to selected promoters in Escherichia coli using bacteriophage lambda and Mu. J Mol Biol. 1976 Jul 5;104(3):541–555. doi: 10.1016/0022-2836(76)90119-4. [DOI] [PubMed] [Google Scholar]
  5. Chappell T. G., Welch W. J., Schlossman D. M., Palter K. B., Schlesinger M. J., Rothman J. E. Uncoating ATPase is a member of the 70 kilodalton family of stress proteins. Cell. 1986 Apr 11;45(1):3–13. doi: 10.1016/0092-8674(86)90532-5. [DOI] [PubMed] [Google Scholar]
  6. Craig E. A., Jacobsen K. Mutations in cognate genes of Saccharomyces cerevisiae hsp70 result in reduced growth rates at low temperatures. Mol Cell Biol. 1985 Dec;5(12):3517–3524. doi: 10.1128/mcb.5.12.3517. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Craig E. A. The heat shock response. CRC Crit Rev Biochem. 1985;18(3):239–280. doi: 10.3109/10409238509085135. [DOI] [PubMed] [Google Scholar]
  8. Echols H. Multiple DNA-protein interactions governing high-precision DNA transactions. Science. 1986 Sep 5;233(4768):1050–1056. doi: 10.1126/science.2943018. [DOI] [PubMed] [Google Scholar]
  9. Ellis J. Proteins as molecular chaperones. 1987 Jul 30-Aug 5Nature. 328(6129):378–379. doi: 10.1038/328378a0. [DOI] [PubMed] [Google Scholar]
  10. Gallant J. A. Stringent control in E. coli. Annu Rev Genet. 1979;13:393–415. doi: 10.1146/annurev.ge.13.120179.002141. [DOI] [PubMed] [Google Scholar]
  11. Georgopoulos C. P., Eisen H. Bacterial mutants which block phage assembly. J Supramol Struct. 1974;2(2-4):349–359. doi: 10.1002/jss.400020224. [DOI] [PubMed] [Google Scholar]
  12. Gomes S. L., Juliani M. H., Maia J. C., Silva A. M. Heat shock protein synthesis during development in Caulobacter crescentus. J Bacteriol. 1986 Nov;168(2):923–930. doi: 10.1128/jb.168.2.923-930.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Grossman A. D., Straus D. B., Walter W. A., Gross C. A. Sigma 32 synthesis can regulate the synthesis of heat shock proteins in Escherichia coli. Genes Dev. 1987 Apr;1(2):179–184. doi: 10.1101/gad.1.2.179. [DOI] [PubMed] [Google Scholar]
  14. 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]
  15. Hemmingsen S. M., Woolford C., van der Vies S. M., Tilly K., Dennis D. T., Georgopoulos C. P., Hendrix R. W., Ellis R. J. Homologous plant and bacterial proteins chaperone oligomeric protein assembly. Nature. 1988 May 26;333(6171):330–334. doi: 10.1038/333330a0. [DOI] [PubMed] [Google Scholar]
  16. Herendeen S. L., VanBogelen R. A., Neidhardt F. C. Levels of major proteins of Escherichia coli during growth at different temperatures. J Bacteriol. 1979 Jul;139(1):185–194. doi: 10.1128/jb.139.1.185-194.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Holland I. B. Genetic analysis of the E. coli division clock. Cell. 1987 Feb 13;48(3):361–362. doi: 10.1016/0092-8674(87)90183-8. [DOI] [PubMed] [Google Scholar]
  18. Horiuchi T., Maki H., Sekiguchi M. RNase H-defective mutants of Escherichia coli: a possible discriminatory role of RNase H in initiation of DNA replication. Mol Gen Genet. 1984;195(1-2):17–22. doi: 10.1007/BF00332717. [DOI] [PubMed] [Google Scholar]
  19. Hunt C., Morimoto R. I. Conserved features of eukaryotic hsp70 genes revealed by comparison with the nucleotide sequence of human hsp70. Proc Natl Acad Sci U S A. 1985 Oct;82(19):6455–6459. doi: 10.1073/pnas.82.19.6455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Itikawa H., Fujita H., Wada M. High temperature induction of a stringent response in the dnaK(Ts) and dnaJ(Ts) mutants of Escherichia coli. J Biochem. 1986 Jun;99(6):1719–1724. doi: 10.1093/oxfordjournals.jbchem.a135648. [DOI] [PubMed] [Google Scholar]
  21. Itikawa H., Ryu J. Isolation and characterization of a temperature-sensitive dnaK mutant of Escherichia coli B. J Bacteriol. 1979 May;138(2):339–344. doi: 10.1128/jb.138.2.339-344.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jones P. G., VanBogelen R. A., Neidhardt F. C. Induction of proteins in response to low temperature in Escherichia coli. J Bacteriol. 1987 May;169(5):2092–2095. doi: 10.1128/jb.169.5.2092-2095.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. KELLENBERGER E., RYTER A., SECHAUD J. Electron microscope study of DNA-containing plasms. II. Vegetative and mature phage DNA as compared with normal bacterial nucleoids in different physiological states. J Biophys Biochem Cytol. 1958 Nov 25;4(6):671–678. doi: 10.1083/jcb.4.6.671. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kao H. T., Capasso O., Heintz N., Nevins J. R. Cell cycle control of the human HSP70 gene: implications for the role of a cellular E1A-like function. Mol Cell Biol. 1985 Apr;5(4):628–633. doi: 10.1128/mcb.5.4.628. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Krueger J. H., Walker G. C. groEL and dnaK genes of Escherichia coli are induced by UV irradiation and nalidixic acid in an htpR+-dependent fashion. Proc Natl Acad Sci U S A. 1984 Mar;81(5):1499–1503. doi: 10.1073/pnas.81.5.1499. [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. Lindquist S. The heat-shock response. Annu Rev Biochem. 1986;55:1151–1191. doi: 10.1146/annurev.bi.55.070186.005443. [DOI] [PubMed] [Google Scholar]
  28. Lutkenhaus J., Sanjanwala B., Lowe M. Overproduction of FtsZ suppresses sensitivity of lon mutants to division inhibition. J Bacteriol. 1986 Jun;166(3):756–762. doi: 10.1128/jb.166.3.756-762.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. McMullin T. W., Hallberg R. L. A highly evolutionarily conserved mitochondrial protein is structurally related to the protein encoded by the Escherichia coli groEL gene. Mol Cell Biol. 1988 Jan;8(1):371–380. doi: 10.1128/mcb.8.1.371. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Milarski K. L., Morimoto R. I. Expression of human HSP70 during the synthetic phase of the cell cycle. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9517–9521. doi: 10.1073/pnas.83.24.9517. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Neidhardt F. C., VanBogelen R. A. Positive regulatory gene for temperature-controlled proteins in Escherichia coli. Biochem Biophys Res Commun. 1981 May 29;100(2):894–900. doi: 10.1016/s0006-291x(81)80257-4. [DOI] [PubMed] [Google Scholar]
  32. Neidhardt F. C., VanBogelen R. A., Vaughn V. The genetics and regulation of heat-shock proteins. Annu Rev Genet. 1984;18:295–329. doi: 10.1146/annurev.ge.18.120184.001455. [DOI] [PubMed] [Google Scholar]
  33. Ogawa T., Pickett G. G., Kogoma T., Kornberg A. RNase H confers specificity in the dnaA-dependent initiation of replication at the unique origin of the Escherichia coli chromosome in vivo and in vitro. Proc Natl Acad Sci U S A. 1984 Feb;81(4):1040–1044. doi: 10.1073/pnas.81.4.1040. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. 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]
  35. Pelham H. R. Speculations on the functions of the major heat shock and glucose-regulated proteins. Cell. 1986 Sep 26;46(7):959–961. doi: 10.1016/0092-8674(86)90693-8. [DOI] [PubMed] [Google Scholar]
  36. Saito H., Uchida H. Initiation of the DNA replication of bacteriophage lambda in Escherichia coli K12. J Mol Biol. 1977 Jun 15;113(1):1–25. doi: 10.1016/0022-2836(77)90038-9. [DOI] [PubMed] [Google Scholar]
  37. Saito H., Uchida H. Organization and expression of the dnaJ and dnaK genes of Escherichia coli K12. Mol Gen Genet. 1978 Aug 4;164(1):1–8. doi: 10.1007/BF00267592. [DOI] [PubMed] [Google Scholar]
  38. Sakakibara Y. The dnaK gene of Escherichia coli functions in initiation of chromosome replication. J Bacteriol. 1988 Feb;170(2):972–979. doi: 10.1128/jb.170.2.972-979.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Sunshine M., Feiss M., Stuart J., Yochem J. A new host gene (groPC) necessary for lambda DNA replication. Mol Gen Genet. 1977 Feb 28;151(1):27–34. doi: 10.1007/BF00446909. [DOI] [PubMed] [Google Scholar]
  40. Tilly K., McKittrick N., Zylicz M., Georgopoulos C. The dnaK protein modulates the heat-shock response of Escherichia coli. Cell. 1983 Sep;34(2):641–646. doi: 10.1016/0092-8674(83)90396-3. [DOI] [PubMed] [Google Scholar]
  41. Tsuchido T., VanBogelen R. A., Neidhardt F. C. Heat shock response in Escherichia coli influences cell division. Proc Natl Acad Sci U S A. 1986 Sep;83(18):6959–6963. doi: 10.1073/pnas.83.18.6959. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Uzan M., Danchin A. A rapid test for the rel A mutation in E. coli. Biochem Biophys Res Commun. 1976 Apr 5;69(3):751–758. doi: 10.1016/0006-291x(76)90939-6. [DOI] [PubMed] [Google Scholar]
  43. Wada M., Sekine K., Itikawa H. Participation of the dnaK and dnaJ gene products in phosphorylation of glutaminyl-tRNA synthetase and threonyl-tRNA synthetase of Escherichia coli K-12. J Bacteriol. 1986 Oct;168(1):213–220. doi: 10.1128/jb.168.1.213-220.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Walker G. C. Mutagenesis and inducible responses to deoxyribonucleic acid damage in Escherichia coli. Microbiol Rev. 1984 Mar;48(1):60–93. doi: 10.1128/mr.48.1.60-93.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Ward J. E., Jr, Lutkenhaus J. Overproduction of FtsZ induces minicell formation in E. coli. Cell. 1985 Oct;42(3):941–949. doi: 10.1016/0092-8674(85)90290-9. [DOI] [PubMed] [Google Scholar]
  46. Winans S. C., Elledge S. J., Krueger J. H., Walker G. C. Site-directed insertion and deletion mutagenesis with cloned fragments in Escherichia coli. J Bacteriol. 1985 Mar;161(3):1219–1221. doi: 10.1128/jb.161.3.1219-1221.1985. [DOI] [PMC free article] [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. Yin J. C., Reznikoff W. S. dnaA, an essential host gene, and Tn5 transposition. J Bacteriol. 1987 Oct;169(10):4637–4645. doi: 10.1128/jb.169.10.4637-4645.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Young D., Lathigra R., Hendrix R., Sweetser D., Young R. A. Stress proteins are immune targets in leprosy and tuberculosis. Proc Natl Acad Sci U S A. 1988 Jun;85(12):4267–4270. doi: 10.1073/pnas.85.12.4267. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. 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]
  51. Zylicz M., Ang D., Georgopoulos C. The grpE protein of Escherichia coli. Purification and properties. J Biol Chem. 1987 Dec 25;262(36):17437–17442. [PubMed] [Google Scholar]
  52. Zylicz M., LeBowitz J. H., McMacken R., Georgopoulos C. The dnaK protein of Escherichia coli possesses an ATPase and autophosphorylating activity and is essential in an in vitro DNA replication system. Proc Natl Acad Sci U S A. 1983 Nov;80(21):6431–6435. doi: 10.1073/pnas.80.21.6431. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES