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. 1986 Nov;6(11):3854–3861. doi: 10.1128/mcb.6.11.3854

Characterization of a Tetrahymena thermophila mutant strain unable to develop normal thermotolerance.

K W Kraus, E M Hallberg, R Hallberg
PMCID: PMC367148  PMID: 3796597

Abstract

For Tetrahymena thermophila cells to survive extended periods of time at 43 degrees C, they must continuously synthesize heat shock proteins. For its translational machinery to function at 43 degrees C, T. thermophila requires either prior nonlethal heat shock treatment or brief treatment with partially inhibiting doses of cycloheximide or emetine. We have identified and characterized a mutant strain of T. thermophila (MC-3) in which prior nonlethal heat shock does not prevent protein synthesis inactivation at 43 degrees C. In addition, treatment of MC-3 cells with either of the antibiotics that normally confer 43 degrees C thermoprotection on wild-type cells elicited no similar thermoprotective response in these cells. Despite these phenotypic characteristics, by other criteria MC-3 synthesized a normal, functional array of heat shock proteins at 40 degrees C, a nonlethal heat shock protein-inducing temperature. The mutation in MC-3 which prevents the thermostabilization of protein synthesis by nonlethal heat shock is, by genetic criteria, most likely the same one which prevents the induction of thermotolerance by drug treatments. We present evidence that this mutation may affect some ribosome-associated functions.

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Selected References

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  1. Ares M., Jr, Bruns P. J. Isolation and genetic characterization of a mutation affecting ribosomal resistance to cycloheximide in Tetrahymena. Genetics. 1978 Nov;90(3):463–474. doi: 10.1093/genetics/90.3.463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bruns P. J., Brussard T. B. Positive selection for mating with functional heterokaryons in Tetrahymena pyriformis. Genetics. 1974 Nov;78(3):831–841. doi: 10.1093/genetics/78.3.831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Calzone F. J., Angerer R. C., Gorovsky M. A. Regulation of protein synthesis in Tetrahymena: isolation and characterization of polysomes by gel filtration and precipitation at pH 5.3. Nucleic Acids Res. 1982 Mar 25;10(6):2145–2161. doi: 10.1093/nar/10.6.2145. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Craig E. A., Jacobsen K. Mutations of the heat inducible 70 kilodalton genes of yeast confer temperature sensitive growth. Cell. 1984 Oct;38(3):841–849. doi: 10.1016/0092-8674(84)90279-4. [DOI] [PubMed] [Google Scholar]
  5. Duncan R., Hershey J. W. Heat shock-induced translational alterations in HeLa cells. Initiation factor modifications and the inhibition of translation. J Biol Chem. 1984 Oct 10;259(19):11882–11889. [PubMed] [Google Scholar]
  6. Gerner E. W., Schneider M. J. Induced thermal resistance in HeLa cells. Nature. 1975 Aug 7;256(5517):500–502. doi: 10.1038/256500a0. [DOI] [PubMed] [Google Scholar]
  7. Guttman S. D., Glover C. V., Allis C. D., Gorovsky M. A. Heat shock, deciliation and release from anoxia induce the synthesis of the same set of polypeptides in starved T. pyriformis. Cell. 1980 Nov;22(1 Pt 1):299–307. doi: 10.1016/0092-8674(80)90177-4. [DOI] [PubMed] [Google Scholar]
  8. Hahn G. M., Li G. C. Thermotolerance and heat shock proteins in mammalian cells. Radiat Res. 1982 Dec;92(3):452–457. [PubMed] [Google Scholar]
  9. Hallberg R. L., Bruns P. J. Ribosome biosynthesis in Tetrahymena pyriformis. Regulation in response to nutritional changes. J Cell Biol. 1976 Nov;71(2):383–394. doi: 10.1083/jcb.71.2.383. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hallberg R. L., Hallberg E. M. Characterization of a cycloheximide-resistant Tetrahymena thermophila mutant which also displays altered growth properties. Mol Cell Biol. 1983 Apr;3(4):503–510. doi: 10.1128/mcb.3.4.503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hallberg R. L., Kraus K. W., Findly R. C. Starved Tetrahymena thermophila cells that are unable to mount an effective heat shock response selectively degrade their rRNA. Mol Cell Biol. 1984 Oct;4(10):2170–2179. doi: 10.1128/mcb.4.10.2170. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hallberg R. L., Kraus K. W., Hallberg E. M. Induction of acquired thermotolerance in Tetrahymena thermophila: effects of protein synthesis inhibitors. Mol Cell Biol. 1985 Aug;5(8):2061–2069. doi: 10.1128/mcb.5.8.2061. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hallberg R. L. No heat shock protein synthesis is required for induced thermostabilization of translational machinery. Mol Cell Biol. 1986 Jun;6(6):2267–2270. doi: 10.1128/mcb.6.6.2267. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hallberg R. L., Wilson P. G., Sutton C. Regulation of ribosome phosphorylation and antibiotic sensitivity in Tetrahymena thermophila: A correlation. Cell. 1981 Oct;26(1 Pt 1):47–56. doi: 10.1016/0092-8674(81)90032-5. [DOI] [PubMed] [Google Scholar]
  15. Henle K. J., Leeper D. B. Interaction of hyperthermia and radiation in CHO cells: recovery kinetics. Radiat Res. 1976 Jun;66(3):505–518. [PubMed] [Google Scholar]
  16. Hickey E. D., Weber L. A. Modulation of heat-shock polypeptide synthesis in HeLa cells during hyperthermia and recovery. Biochemistry. 1982 Mar 30;21(7):1513–1521. doi: 10.1021/bi00536a008. [DOI] [PubMed] [Google Scholar]
  17. Iida H., Yahara I. A heat shock-resistant mutant of Saccharomyces cerevisiae shows constitutive synthesis of two heat shock proteins and altered growth. J Cell Biol. 1984 Oct;99(4 Pt 1):1441–1450. doi: 10.1083/jcb.99.4.1441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Krüger C., Benecke B. J. In vitro translation of Drosophila heat-shock and non--heat-shock mRNAs in heterologous and homologous cell-free systems. Cell. 1981 Feb;23(2):595–603. doi: 10.1016/0092-8674(81)90155-0. [DOI] [PubMed] [Google Scholar]
  19. Li G. C. Induction of thermotolerance and enhanced heat shock protein synthesis in Chinese hamster fibroblasts by sodium arsenite and by ethanol. J Cell Physiol. 1983 May;115(2):116–122. doi: 10.1002/jcp.1041150203. [DOI] [PubMed] [Google Scholar]
  20. Li G. C., Werb Z. Correlation between synthesis of heat shock proteins and development of thermotolerance in Chinese hamster fibroblasts. Proc Natl Acad Sci U S A. 1982 May;79(10):3218–3222. doi: 10.1073/pnas.79.10.3218. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lin C. Y., Roberts J. K., Key J. L. Acquisition of Thermotolerance in Soybean Seedlings : Synthesis and Accumulation of Heat Shock Proteins and their Cellular Localization. Plant Physiol. 1984 Jan;74(1):152–160. doi: 10.1104/pp.74.1.152. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Loomis W. F., Wheeler S. A. Chromatin-associated heat shock proteins of Dictyostelium. Dev Biol. 1982 Apr;90(2):412–418. doi: 10.1016/0012-1606(82)90390-6. [DOI] [PubMed] [Google Scholar]
  23. Martindale D. W., Bruns P. J. Cloning of abundant mRNA species present during conjugation of Tetrahymena thermophila: identification of mRNA species present exclusively during meiosis. Mol Cell Biol. 1983 Oct;3(10):1857–1865. doi: 10.1128/mcb.3.10.1857. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. McMaster G. K., Carmichael G. G. Analysis of single- and double-stranded nucleic acids on polyacrylamide and agarose gels by using glyoxal and acridine orange. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4835–4838. doi: 10.1073/pnas.74.11.4835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. McMullin T. W., Hallberg R. L. Effect of heat shock on ribosome structure: appearance of a new ribosome-associated protein. Mol Cell Biol. 1986 Jul;6(7):2527–2535. doi: 10.1128/mcb.6.7.2527. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Orias E., Bruns P. J. Induction and isolation of mutants in Tetrahymena. Methods Cell Biol. 1976;13:247–282. [PubMed] [Google Scholar]
  27. Orias E., Hamilton E. P. Cytogamy: An Inducible, Alternate Pathway of Conjugation in TETRAHYMENA THERMOPHILA. Genetics. 1979 Apr;91(4):657–671. doi: 10.1093/genetics/91.4.657. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Petersen N. S., Mitchell H. K. Recovery of protein synthesis after heat shock: prior heat treatment affects the ability of cells to translate mRNA. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1708–1711. doi: 10.1073/pnas.78.3.1708. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Roberts C. T., Jr, Orias E. On the mechanism of adaptation to protein synthesis inhibitors by Tetrahymena. Facilitation, cross adaptation, and resensitization. J Cell Biol. 1974 Sep;62(3):707–716. doi: 10.1083/jcb.62.3.707. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Scharf K. D., Nover L. Heat-shock-induced alterations of ribosomal protein phosphorylation in plant cell cultures. Cell. 1982 Sep;30(2):427–437. doi: 10.1016/0092-8674(82)90240-9. [DOI] [PubMed] [Google Scholar]
  31. Scott M. P., Pardue M. L. Translational control in lysates of Drosophila melanogaster cells. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3353–3357. doi: 10.1073/pnas.78.6.3353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Storti R. V., Scott M. P., Rich A., Pardue M. L. Translational control of protein synthesis in response to heat shock in D. melanogaster cells. Cell. 1980 Dec;22(3):825–834. doi: 10.1016/0092-8674(80)90559-0. [DOI] [PubMed] [Google Scholar]
  33. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Wang T. C., Hooper A. B. Adaptation to cycloheximide of macromolecular synthesis in Tetrahymena. J Cell Physiol. 1978 Apr;95(1):1–11. doi: 10.1002/jcp.1040950102. [DOI] [PubMed] [Google Scholar]
  35. White B. A., Bancroft F. C. Cytoplasmic dot hybridization. Simple analysis of relative mRNA levels in multiple small cell or tissue samples. J Biol Chem. 1982 Aug 10;257(15):8569–8572. [PubMed] [Google Scholar]

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