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Title: A combinatorial approach to synthetic transcription factor-promoter combinations for yeast strain engineering

Abstract

Despite the need for inducible promoters in strain development efforts, the majority of engineering in Saccharomyces cerevisiae continues to rely on a few constitutively active or inducible promoters. Building on advances that use the modular nature of both transcription factors and promoter regions, we have built a library of hybrid promoters that are regulated by a synthetic transcription factor. The hybrid promoters consist of native S. cerevisiae promoters, in which the operator regions have been replaced with sequences that are recognized by the bacterial LexA DNA binding protein. Correspondingly, the synthetic transcription factor (TF) consists of the DNA binding domain of the LexA protein, fused with the human estrogen binding domain and the viral activator domain, VP16. The resulting system with a bacterial DNA binding domain avoids the transcription of native S. cerevisiae genes, and the hybrid promoters can be induced using estradiol, a compound with no detectable impact on S. cerevisiae physiology. Using combinations of one, two or three operator sequence repeats and a set of native S. cerevisiae promoters, we obtained a series of hybrid promoters that can be induced to different levels, using the same synthetic TF and a given estradiol. Finally, this set of promoters, inmore » combination with our synthetic TF, has the potential to regulate numerous genes or pathways simultaneously, to multiple desired levels, in a single strain.« less

Authors:
 [1];  [1];  [1];  [2];  [3];  [4]; ORCiD logo [1]
  1. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Biological Systems and Engineering Division
  2. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Biological Systems and Engineering Division; USDOE Joint Genome Institute (JGI), Walnut Creek, CA (United States); USDOE Agile BioFoundry, Emeryville, CA (United States)
  3. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Biological Systems and Engineering Division; USDOE Joint Genome Institute (JGI), Walnut Creek, CA (United States); USDOE Agile BioFoundry, Emeryville, CA (United States)
  4. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Biological Systems and Engineering Division; Univ. of California, Berkeley, CA (United States). Dept. of Chemical and Biomolecular Engineering and Dept. of Bioengineering; Technical Univ. of Denmark, Lyngby (Denmark). Novo Nordisk Foundation Center forBiosustainability
Publication Date:
Research Org.:
Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
OSTI Identifier:
1433124
Grant/Contract Number:  
AC02-05CH11231
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Yeast
Additional Journal Information:
Journal Volume: 35; Journal Issue: 3; Journal ID: ISSN 0749-503X
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; hybrid promoter; Saccharomyces; strain engineering; synthetic biology

Citation Formats

Dossani, Zain Y., Reider Apel, Amanda, Szmidt-Middleton, Heather, Hillson, Nathan J., Deutsch, Samuel, Keasling, Jay D., and Mukhopadhyay, Aindrila. A combinatorial approach to synthetic transcription factor-promoter combinations for yeast strain engineering. United States: N. p., 2017. Web. doi:10.1002/yea.3292.
Dossani, Zain Y., Reider Apel, Amanda, Szmidt-Middleton, Heather, Hillson, Nathan J., Deutsch, Samuel, Keasling, Jay D., & Mukhopadhyay, Aindrila. A combinatorial approach to synthetic transcription factor-promoter combinations for yeast strain engineering. United States. doi:10.1002/yea.3292.
Dossani, Zain Y., Reider Apel, Amanda, Szmidt-Middleton, Heather, Hillson, Nathan J., Deutsch, Samuel, Keasling, Jay D., and Mukhopadhyay, Aindrila. Mon . "A combinatorial approach to synthetic transcription factor-promoter combinations for yeast strain engineering". United States. doi:10.1002/yea.3292. https://www.osti.gov/servlets/purl/1433124.
@article{osti_1433124,
title = {A combinatorial approach to synthetic transcription factor-promoter combinations for yeast strain engineering},
author = {Dossani, Zain Y. and Reider Apel, Amanda and Szmidt-Middleton, Heather and Hillson, Nathan J. and Deutsch, Samuel and Keasling, Jay D. and Mukhopadhyay, Aindrila},
abstractNote = {Despite the need for inducible promoters in strain development efforts, the majority of engineering in Saccharomyces cerevisiae continues to rely on a few constitutively active or inducible promoters. Building on advances that use the modular nature of both transcription factors and promoter regions, we have built a library of hybrid promoters that are regulated by a synthetic transcription factor. The hybrid promoters consist of native S. cerevisiae promoters, in which the operator regions have been replaced with sequences that are recognized by the bacterial LexA DNA binding protein. Correspondingly, the synthetic transcription factor (TF) consists of the DNA binding domain of the LexA protein, fused with the human estrogen binding domain and the viral activator domain, VP16. The resulting system with a bacterial DNA binding domain avoids the transcription of native S. cerevisiae genes, and the hybrid promoters can be induced using estradiol, a compound with no detectable impact on S. cerevisiae physiology. Using combinations of one, two or three operator sequence repeats and a set of native S. cerevisiae promoters, we obtained a series of hybrid promoters that can be induced to different levels, using the same synthetic TF and a given estradiol. Finally, this set of promoters, in combination with our synthetic TF, has the potential to regulate numerous genes or pathways simultaneously, to multiple desired levels, in a single strain.},
doi = {10.1002/yea.3292},
journal = {Yeast},
number = 3,
volume = 35,
place = {United States},
year = {Mon Oct 30 00:00:00 EDT 2017},
month = {Mon Oct 30 00:00:00 EDT 2017}
}

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Works referenced in this record:

CRISPR-Mediated Modular RNA-Guided Regulation of Transcription in Eukaryotes
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j5 DNA Assembly Design Automation Software
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