Perturbation-based analysis and modeling of combinatorial regulation in the yeast sulfur assimilation pathway.
In yeast, the pathways of sulfur assimilation are combinatorially controlled by five transcriptional regulators (three DNA-binding proteins [Met31p, Met32p, and Cbf1p], an activator [Met4p], and a cofactor [Met28p]) and a ubiquitin ligase subunit (Met30p). This regulatory system exerts combinatorial control not only over sulfur assimilation and methionine biosynthesis, but also on many other physiological functions in the cell. Recently we characterized a gene induction system that, upon the addition of an inducer, results in near-immediate transcription of a gene of interest under physiological conditions. We used this to perturb levels of single transcription factors during steady-state growth in chemostats, which facilitated distinction of direct from indirect effects of individual factors dynamically through quantification of the subsequent changes in genome-wide patterns of gene expression. We were able to show directly that Cbf1p acts sometimes as a repressor and sometimes as an activator. We also found circumstances in which Met31p/Met32p function as repressors, as well as those in which they function as activators. We elucidated and numerically modeled feedback relationships among the regulators, notably feedforward regulation of Met32p (but not Met31p) by Met4p that generates dynamic differences in abundance that can account for the differences in function of these two proteins despite their identical binding sites.
The Lewis-Sigler Institute for Integrative Genomics, Princeton University, Princeton, NJ 08544, USA. rmcisaac@princeton.edu
R Scott
RS
McIsaac
Allegra A
AA
Petti
Harmen J
HJ
Bussemaker
David
D
Botstein
D013766:5-Methyltetrahydrofolate-Homocysteine S-Methyltransferase / Q000172:deficiency; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000235:genetics / Q000502:physiology; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology; D044783:F-Box Proteins / Q000235:genetics / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D016681:Genome, Fungal; D008715:Methionine / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D013455:Sulfur / Q000378:metabolism* / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology; D014158:Transcription, Genetic; D025801:Ubiquitin / Q000378:metabolism; D043743:Ubiquitin-Protein Ligase Complexes / Q000235:genetics / Q000378:metabolism
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; C064042:CBF1 protein, S cerevisiae; D004268:DNA-Binding Proteins; D044783:F-Box Proteins; C470285:MET30 protein, S cerevisiae; C106764:Met31 protein, S cerevisiae; C106765:Met32 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D025801:Ubiquitin; D013455:Sulfur; D008715:Methionine; D013766:5-Methyltetrahydrofolate-Homocysteine S-Methyltransferase; D043743:Ubiquitin-Protein Ligase Complexes