[29] Superimposition of temperature regulation on yeast promoters

Sledziewski, A; Bell, Anne; Yip, Carli; Kelsay, Kimberly; Grant, Francis James; MacKay, Vivian L.

doi:10.1016/0076-6879(90)85031-i

Cited by 9 publications

(5 citation statements)

References 20 publications

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“…Different from the previously developed temperature‐induced protein expression systems in yeast, which was based on mutation of the acid phosphatase regulatory genes pho80 and pho4 ts (Kramer, DeChiara, Schaber, & Hilliker, ) or mating type control involving sir3 mutation and MATα2‐ hybrid promoters (Sledziewski et al, ), the temperature‐responsive regulation device presented in the current work was modified from the GAL system which has rapid transcriptional enhancement of >1,000‐fold upon inducer addition (Lohr, Venkov, & Zlatanova, ) and wide applications in metabolic regulation, and the temperature sensitivity was acquired by Gal4 directed evolution. Since the structure information of Gal4 is only available for the DNA binding domain, the scope of engineering targets in rational design is largely restricted.…”

Section: Discussionmentioning

confidence: 99%

Development of a temperature‐responsive yeast cell factory using engineered Gal4 as a protein switch

Zhou

Xie

Yao

et al. 2018

Biotech & Bioengineering

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Conflict between cell growth and product accumulation is frequently encountered in biosynthesis of secondary metabolites. Herein, a temperature-dependent dynamic control strategy was developed by modifying the GAL regulation system to facilitate two-stage fermentation in yeast. A temperature-sensitive Gal4 mutant Gal4M9 was created by directed evolution, and used as a protein switch in ΔGAL80 yeast. After EGFP-reported validation of its temperature-responsive induction capability, the sensitivity and stringency of this system in multi-gene pathway regulation was tested, using lycopene as an example product. When Gal4M9 was used to control the expression of P -driven pathway genes, growth and production was successfully decoupled upon temperature shift during fermentation, accumulating 44% higher biomass and 177% more lycopene than the control strain with wild-type Gal4. This is the first example of adopting temperature as an input signal for metabolic pathway regulation in yeast cell factories.

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Section: Discussionmentioning

confidence: 99%

Development of a temperature‐responsive yeast cell factory using engineered Gal4 as a protein switch

Zhou

Xie

Yao

et al. 2018

Biotech & Bioengineering

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“…However, at the restrictive temperature of 35 °C, lack of functional Sir3 protein allows expression of α2 repressor from the HML α cassette, which can repress the target gene. The advantages of such a temperature‐regulated expression system are two‐fold: first, the reproducibility of a transfer of technology to the commercial level in such a system is better, and second, it allows fine tuning or calibration of the expression level of proteins by varying the temperature, as in a rheostat, thus allowing optimization of expression level, which may vary from one protein to another (Sledziewski et al , 1990). The short induction period of 40 min, with maximum levels being achieved after 3 h and a decline after 9 h, may especially be more advantageous than nmt1 / 41 / 81 in executing physiological experiments where a fast induction to monitor the effect of expressing or absence of protein need to be investigated.…”

Section: Discussionmentioning

confidence: 99%

A truncated derivative of nmt1 promoter exhibits temperature‐dependent induction of gene expression in Schizosaccharomyces pombe

Kumar

Singh

2006

Yeast

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Despite increasing exploitation of Schizosaccharomyces pombe as a model system there is a lack of convenient vectors for research and application. Expression with the commonly used promoter, nmt1, requires a laborious regime involving the removal of repressor, thiamine, from a growing culture and further growth for 18 h to achieve maximum expression, thus underlining the need for more user-friendly promoters. We report here the isolation and characterization of a truncated derivative of the nmt1 promoter having novel induction characteristics: it is induced by shift of growth temperature from 36 • C to 25 • C, achieving maximum expression within 3 h. Similar features of expression were observed with the reporter genes GFP and β-galactosidase, a native gene, cdc18, and a commercially important foreign therapeutic protein, streptokinase. The new promoter element offers additional advantages, such as lack of deleterious effect on cell viability and potential ability to express toxic proteins. These features make the new promoter a potentially better alternative to nmt1, both as a research tool and for expression of commercially important proteins in Sz. pombe, and suggest the possibility of using similar approaches to design promoters with novel and useful properties.

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“…In the temperature-sensitive sir3 mutants, expression of the MAT α2 repressor is temperature dependent. Therefore, specifi c target genes cloned downstream of a promoter with this 31 bp UAS sequence are repressed at 35 °C due to the inactivation of Sir3 allowing for the expression of the Mat α 2 repressor [ 55 ]. At the permissive temperature, 25 °C, Sir3 is active and therefore inhibits expression of the Mat α 2 repressor, hence enabling the expression of genes controlled by the Mat α 2 system [ 55 , 57 , 58 ].…”

Section: Regulated Promotersmentioning

confidence: 99%

“…Certain physical stimuli are attractive for implementation in large-scale production processes, particularly if they do not require sophisticated equipment. A temperature-regulated promoter system was developed based on artifi cial thermosensitivity of mating-type regulation in S. cerevisiae [ 55 ]. Specifi cally, a temperature-sensitive mutation in the SIR3 gene renders the silencing protein Sir3 inactive at 35 °C and active at 25 °C.…”

Section: Regulated Promotersmentioning

confidence: 99%

Natural and Modified Promoters for Tailored Metabolic Engineering of the Yeast Saccharomyces cerevisiae

Hubmann

Thevelein

Nevoigt

2014

Methods in Molecular Biology

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The ease of highly sophisticated genetic manipulations in the yeast Saccharomyces cerevisiae has initiated numerous initiatives towards development of metabolically engineered strains for novel applications beyond its traditional use in brewing, baking, and wine making. In fact, baker's yeast has become a key cell factory for the production of various bulk and fine chemicals. Successful metabolic engineering requires fine-tuned adjustments of metabolic fluxes and coordination of multiple pathways within the cell. This has mostly been achieved by controlling gene expression at the transcriptional level, i.e., by using promoters with appropriate strengths and regulatory properties. Here we present an overview of natural and modified promoters, which have been used in metabolic pathway engineering of S. cerevisiae. Recent developments in creating promoters with tailor-made properties are also discussed.

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[29] Superimposition of temperature regulation on yeast promoters

Cited by 9 publications

References 20 publications

Development of a temperature‐responsive yeast cell factory using engineered Gal4 as a protein switch

Development of a temperature‐responsive yeast cell factory using engineered Gal4 as a protein switch

A truncated derivative of nmt1 promoter exhibits temperature‐dependent induction of gene expression in Schizosaccharomyces pombe

Natural and Modified Promoters for Tailored Metabolic Engineering of the Yeast Saccharomyces cerevisiae

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