Fast-acting and nearly gratuitous induction of gene expression and protein depletion in Saccharomyces cerevisiae

R. Scott McIsaac, Sanford J. Silverman, Megan N. McClean, Patrick A. Gibney, Joanna Macinskas, Mark J. Hickman, Allegra A. Petti, David Botstein

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69 Scopus citations

Abstract

We describe the development and characterization of a system that allows the rapid and specific induction of individual genes in the yeast Saccharomyces cerevisiae without changes in nutrients or temperature. The system is based on the chimeric transcriptional activator Gal4dbd.ER.VP16 (GEV). Upon addition of the hormone β-estradiol, cytoplasmic GEV localizes to the nucleus and binds to promoters containing Gal4p consensus binding sequences to activate transcription. With galactokinase Gal1p and transcriptional activator Gal4p absent, the system is fast-acting, resulting in readily detectable transcription within 5 min after addition of the inducer. β-Estradiol is nearly a gratuitous inducer, as indicated by genome-wide profiling that shows unintended induction (by GEV) of only a few dozen genes. Response to inducer is graded: intermediate concentrations of inducer result in production of intermediate levels of product protein in all cells. We present data illustrating several applications of this system, including a modification of the regulated degron method, which allows rapid and specific degradation of a specific protein upon addition of β-estradiol. These gene induction and protein degradation systems provide important tools for studying the dynamics and functional relationships of genes and their respective regulatory networks.

Original languageEnglish
Pages (from-to)4447-4459
Number of pages13
JournalMolecular biology of the cell
Volume22
Issue number22
DOIs
StatePublished - Nov 15 2011

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    McIsaac, R. S., Silverman, S. J., McClean, M. N., Gibney, P. A., Macinskas, J., Hickman, M. J., Petti, A. A., & Botstein, D. (2011). Fast-acting and nearly gratuitous induction of gene expression and protein depletion in Saccharomyces cerevisiae. Molecular biology of the cell, 22(22), 4447-4459. https://doi.org/10.1091/mbc.E11-05-0466