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Studies of the heat shock response of the yeast Saccharomyces cerevisiae

Cheng, Lili; (1994) Studies of the heat shock response of the yeast Saccharomyces cerevisiae. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

Heat shock protein HSP90 has recently been found to be one of the major chaperones of eukaryotic cells. Saccharomyces cerevisiae transformants were constructed with 50-150 copies of the homologous heat-inducible gene for HSP90 (HSP82) present on a high copy number episomal vector. These transformants were then used to demonstrate: (i) that this gene, normally single-copy in the haploid yeast genome, displays almost normal basal and heat shock-induced levels of expression even at 50-150 copies per cell; (ii) that yeast is an expression host suitable for high level synthesis of HSP90 protein (to 30-40% of total cell protein); and (iii) that increasing cellular levels of HSP90 is not protective against heat killing, causing strain-dependent reductions in growth at 37.5°C and in thermotolerance. Heat shock protein induction is only one change elicited in yeast by heat shock. Trehalose is also accumulated, while declining intracellular pH stimulates plasma membrane ATPase activity. Recently the trehalose induction was shown to be regulated by levels of HSP70 and, to a lesser extent, HSP104. Another HSP which might contribute to regulation is HSP90, especially as HSP90 forms complexes with heat shock transcription factor and several of the regulatory proteins of eukaryotic cells. This possibility was investigated using isogenic yeast strains with normal, decreased or elevated HSP90. The results show HSP90 levels having a small negative influence over the heat inductions of trehalose and the heat shock element, a minor effect compared with the major regulation exerted by HSP70. Numerous chemicals are inducers of heat shock proteins, but few agents are known that selectively inhibit expression of these proteins. This thesis provides the first evidence that two widely-used food preservatives (sorbate and benzoate) can act as selective inhibitors of heat shock protein induction in S. cerevisiae. Their effects were strongly dependent on the pH of the culture medium. Below pH5.5 sorbate caused a greatly- increased sensitivity to lethal heat treatment, with strong selection for respiratory- deficient rho- petites amongst the surviving cells. In contrast above pH5.5 sorbate acted as a chemical inducer of thermotolerance. In low pH rho+ cultures sorbate, benzoate, the uncoupler carbonyl cyanide m-chlorophenylhydrazone and the plasma membrane ATPase inhibitor diethylstilboestrol were all shown to act as selective inhibitors of heat shock protein and a heat shock element-lacZ fusion induction by heat shock. In low pH cultures the rho- mutation was found to confer a higher resistance to growth in the presence of sorbate, as well as a partial restoration of the heat inducibility of heat shock proteins and of heat shock element-lacZ fusion in sorbate-treated cultures.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Studies of the heat shock response of the yeast Saccharomyces cerevisiae
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
Keywords: Biological sciences; Heat shock protein
URI: https://discovery.ucl.ac.uk/id/eprint/10107008
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