• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of molcellbPermissionsJournals.ASM.orgJournalMCB ArticleJournal InfoAuthorsReviewers
Mol Cell Biol. Sep 1983; 3(9): 1625–1633.
PMCID: PMC370016

Heat shock-regulated production of Escherichia coli beta-galactosidase in Saccharomyces cerevisiae.

Abstract

The HSP90 gene of the yeast Saccharomyces cerevisiae encodes a heat shock-inducible protein with an Mr of 90,000 (hsp90) and unknown function. We fused DNA fragments of a known sequence (namely, either end of a 1.4-kilobase EcoRI fragment which contains the S. cerevisiae TRP1 gene) to an EcoRI site within the coding sequence of the HSP90 gene. When these fusions are introduced into S. cerevisiae they direct the synthesis of unique truncated hsp90 proteins. By determining the size and charge of these proteins we were able to deduce the translational reading frame at the (EcoRI) fusion site. This information allowed us to design and construct a well-defined in-frame fusion between the S. cerevisiae HSP90 gene and the Escherichia coli lacZ gene. When this fused gene is introduced into S. cerevisiae on a multicopy plasmid vector, it directs the heat shock-inducible synthesis of a fused protein, which is an enzymatically active beta-galactosidase. Thus, for the first time, it is possible to quantitate the heat shock response in a eucaryotic organism with a simple enzyme assay.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.6M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Ashburner M, Bonner JJ. The induction of gene activity in drosophilia by heat shock. Cell. 1979 Jun;17(2):241–254. [PubMed]
  • Beggs JD. Transformation of yeast by a replicating hybrid plasmid. Nature. 1978 Sep 14;275(5676):104–109. [PubMed]
  • Bennetzen JL, Hall BD. Codon selection in yeast. J Biol Chem. 1982 Mar 25;257(6):3026–3031. [PubMed]
  • Broach JR, Hicks JB. Replication and recombination functions associated with the yeast plasmid, 2 mu circle. Cell. 1980 Sep;21(2):501–508. [PubMed]
  • Burnette WN. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. [PubMed]
  • Casadaban MJ, Chou J, Cohen SN. In vitro gene fusions that join an enzymatically active beta-galactosidase segment to amino-terminal fragments of exogenous proteins: Escherichia coli plasmid vectors for the detection and cloning of translational initiation signals. J Bacteriol. 1980 Aug;143(2):971–980. [PMC free article] [PubMed]
  • Corces V, Pellicer A, Axel R, Meselson M. Integration, transcription, and control of a Drosophila heat shock gene in mouse cells. Proc Natl Acad Sci U S A. 1981 Nov;78(11):7038–7042. [PMC free article] [PubMed]
  • Finkelstein DB, Butow RA. DNA-binding proteins in yeast. Effect of growth phase and mitochondrial function. Arch Biochem Biophys. 1976 May;174(1):52–65. [PubMed]
  • Finkelstein DB, Strausberg S. Identification and expression of a cloned yeast heat shock gene. J Biol Chem. 1983 Feb 10;258(3):1908–1913. [PubMed]
  • Finkelstein DB, Strausberg S, McAlister L. Alterations of transcription during heat shock of Saccharomyces cerevisiae. J Biol Chem. 1982 Jul 25;257(14):8405–8411. [PubMed]
  • Guarente L, Ptashne M. Fusion of Escherichia coli lacZ to the cytochrome c gene of Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2199–2203. [PMC free article] [PubMed]
  • Rose M, Casadaban MJ, Botstein D. Yeast genes fused to beta-galactosidase in Escherichia coli can be expressed normally in yeast. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2460–2464. [PMC free article] [PubMed]
  • Stinchcomb DT, Struhl K, Davis RW. Isolation and characterisation of a yeast chromosomal replicator. Nature. 1979 Nov 1;282(5734):39–43. [PubMed]
  • Tschumper G, Carbon J. Sequence of a yeast DNA fragment containing a chromosomal replicator and the TRP1 gene. Gene. 1980 Jul;10(2):157–166. [PubMed]
  • Yamamori T, Ito K, Nakamura Y, Yura T. Transient regulation of protein synthesis in Escherichia coli upon shift-up of growth temperature. J Bacteriol. 1978 Jun;134(3):1133–1140. [PMC free article] [PubMed]
  • Zaret KS, Sherman F. DNA sequence required for efficient transcription termination in yeast. Cell. 1982 Mar;28(3):563–573. [PubMed]

Articles from Molecular and Cellular Biology are provided here courtesy of American Society for Microbiology (ASM)

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...