Rapamycin conditionally inhibits Hsp90 but not Hsp70 mRNA translation in Drosophila: implications for the mechanisms of Hsp mRNA translation

Abstract

Rapamycin inhibits the activity of the target of rapamycin (TOR)-dependent signaling pathway, which has been characterized as one dedicated to translational regulation through modulating cap-dependent translation, involving eIF4E binding protein (eIF4E-BP) or 4E-BP. Results show that rapamycin strongly inhibits global translation in Drosophila cells. However, Hsp70 mRNA translation is virtually unaffected by rapamycin treatment, whereas Hsp90 mRNA translation is strongly inhibited, at normal growth temperature. Intriguingly, during heat shock Hsp90 mRNA becomes significantly less sensitive to rapamycin-mediated inhibition, suggesting the pathway for Hsp90 mRNA translation is altered during heat shock. Reporter mRNAs containing the Hsp90 or Hsp70 mRNAs’ 5′ untranslated region recapitulate these rapamycin-dependent translational characteristics, indicating this region regulates rapamycin-dependent translational sensitivity as well as heat shock preferential translation. Surprisingly, rapamycin-mediated inhibition of Hsp90 mRNA translation at normal growth temperature is not caused by 4E-BP-mediated inhibition of cap-dependent translation. Indeed, no evidence for rapamycin-mediated impaired eIF4E function is observed. These results support the proposal that preferential translation of different Hsp mRNA utilizes distinct translation mechanisms, even within a single species.

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Abbreviations

TOR:

Target of rapamycin

S6K:

ribosomal protein S6 kinase

PI3K:

phosphatidyl inositol 3-kinase

eIF:

eukaryotic initiation factor

4E-BP:

eIF4E binding protein (eIF4E-BP)

5′UTR:

5′ untranslated region of mRNA

FBS:

fetal bovine serum

SM:

Schneider's medium

EBSS:

Earle's buffered salt solution

PABP or dmPABP:

poly(A)-binding protein (from Drosophila melanogaster)

“d” prefix precedingmolecular designations:

Drosophila forms of the enzymes

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Acknowledgements

This research was supported by NSF grant MCB-9728753. We thank Drs. N. Sonenberg, M. Miron, and J. Sierra for providing antisera to Drosophila eIF4E, 4E-BP, and eIF4G. We thank Dr. A. Vincent for providing antisera to Drosophila PABP. We thank Dr. R. Ahmed for the preparation of the plasmid vectors.

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Correspondence to Roger F. Duncan.

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Duncan, R.F. Rapamycin conditionally inhibits Hsp90 but not Hsp70 mRNA translation in Drosophila: implications for the mechanisms of Hsp mRNA translation. Cell Stress and Chaperones 13, 143–155 (2008). https://doi.org/10.1007/s12192-008-0024-6

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Keywords

  • Translation
  • Heat shock
  • Rapamycin
  • Hsp90
  • Hsp70