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Tailor-Made Zinc-Finger Transcription Factors Activate FLO11 Gene Expression with Phenotypic Consequences in the Yeast Saccharomyces cerevisiae

Cys(2)His(2) zinc fingers are eukaryotic DNA-binding motifs, capable of distinguishing different DNA sequences, and are suitable for engineering artificial transcription factors. In this work, we used the budding yeast Saccharomyces cerevisiae to study the ability of tailor-made zinc finger proteins...

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Autores principales: Shieh, Jia-Ching, Cheng, Yu-Che, Su, Mao-Chang, Moore, Michael, Choo, Yen, Klug, Aaron
Formato: Texto
Lenguaje:English
Publicado: Public Library of Science 2007
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1939876/
https://www.ncbi.nlm.nih.gov/pubmed/17710146
http://dx.doi.org/10.1371/journal.pone.0000746
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author Shieh, Jia-Ching
Cheng, Yu-Che
Su, Mao-Chang
Moore, Michael
Choo, Yen
Klug, Aaron
author_facet Shieh, Jia-Ching
Cheng, Yu-Che
Su, Mao-Chang
Moore, Michael
Choo, Yen
Klug, Aaron
author_sort Shieh, Jia-Ching
collection PubMed
description Cys(2)His(2) zinc fingers are eukaryotic DNA-binding motifs, capable of distinguishing different DNA sequences, and are suitable for engineering artificial transcription factors. In this work, we used the budding yeast Saccharomyces cerevisiae to study the ability of tailor-made zinc finger proteins to activate the expression of the FLO11 gene, with phenotypic consequences. Two three-finger peptides were identified, recognizing sites from the 5′ UTR of the FLO11 gene with nanomolar DNA-binding affinity. The three-finger domains and their combined six-finger motif, recognizing an 18-bp site, were fused to the activation domain of VP16 or VP64. These transcription factor constructs retained their DNA-binding ability, with the six-finger ones being the highest in affinity. However, when expressed in haploid yeast cells, only one three-finger recombinant transcription factor was able to activate the expression of FLO11 efficiently. Unlike in the wild-type, cells with such transcriptional activation displayed invasive growth and biofilm formation, without any requirement for glucose depletion. The VP16 and VP64 domains appeared to act equally well in the activation of FLO11 expression, with comparable effects in phenotypic alteration. We conclude that the functional activity of tailor-made transcription factors in cells is not easily predicted by the in vitro DNA-binding activity.
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spelling pubmed-19398762007-08-15 Tailor-Made Zinc-Finger Transcription Factors Activate FLO11 Gene Expression with Phenotypic Consequences in the Yeast Saccharomyces cerevisiae Shieh, Jia-Ching Cheng, Yu-Che Su, Mao-Chang Moore, Michael Choo, Yen Klug, Aaron PLoS One Research Article Cys(2)His(2) zinc fingers are eukaryotic DNA-binding motifs, capable of distinguishing different DNA sequences, and are suitable for engineering artificial transcription factors. In this work, we used the budding yeast Saccharomyces cerevisiae to study the ability of tailor-made zinc finger proteins to activate the expression of the FLO11 gene, with phenotypic consequences. Two three-finger peptides were identified, recognizing sites from the 5′ UTR of the FLO11 gene with nanomolar DNA-binding affinity. The three-finger domains and their combined six-finger motif, recognizing an 18-bp site, were fused to the activation domain of VP16 or VP64. These transcription factor constructs retained their DNA-binding ability, with the six-finger ones being the highest in affinity. However, when expressed in haploid yeast cells, only one three-finger recombinant transcription factor was able to activate the expression of FLO11 efficiently. Unlike in the wild-type, cells with such transcriptional activation displayed invasive growth and biofilm formation, without any requirement for glucose depletion. The VP16 and VP64 domains appeared to act equally well in the activation of FLO11 expression, with comparable effects in phenotypic alteration. We conclude that the functional activity of tailor-made transcription factors in cells is not easily predicted by the in vitro DNA-binding activity. Public Library of Science 2007-08-15 /pmc/articles/PMC1939876/ /pubmed/17710146 http://dx.doi.org/10.1371/journal.pone.0000746 Text en Shieh et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Shieh, Jia-Ching
Cheng, Yu-Che
Su, Mao-Chang
Moore, Michael
Choo, Yen
Klug, Aaron
Tailor-Made Zinc-Finger Transcription Factors Activate FLO11 Gene Expression with Phenotypic Consequences in the Yeast Saccharomyces cerevisiae
title Tailor-Made Zinc-Finger Transcription Factors Activate FLO11 Gene Expression with Phenotypic Consequences in the Yeast Saccharomyces cerevisiae
title_full Tailor-Made Zinc-Finger Transcription Factors Activate FLO11 Gene Expression with Phenotypic Consequences in the Yeast Saccharomyces cerevisiae
title_fullStr Tailor-Made Zinc-Finger Transcription Factors Activate FLO11 Gene Expression with Phenotypic Consequences in the Yeast Saccharomyces cerevisiae
title_full_unstemmed Tailor-Made Zinc-Finger Transcription Factors Activate FLO11 Gene Expression with Phenotypic Consequences in the Yeast Saccharomyces cerevisiae
title_short Tailor-Made Zinc-Finger Transcription Factors Activate FLO11 Gene Expression with Phenotypic Consequences in the Yeast Saccharomyces cerevisiae
title_sort tailor-made zinc-finger transcription factors activate flo11 gene expression with phenotypic consequences in the yeast saccharomyces cerevisiae
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1939876/
https://www.ncbi.nlm.nih.gov/pubmed/17710146
http://dx.doi.org/10.1371/journal.pone.0000746
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