Determinants of the ubiquitin-mediated degradation of the Met4 transcription factor.
In yeast, the Met4 transcription factor and its cofactors Cbf1, Met28, Met31, and Met32 control the expression of sulfur metabolism and oxidative stress response genes. Met4 activity is tuned to nutrient and oxidative stress conditions by the SCF(Met30) ubiquitin ligase. The mechanism whereby SCF(Met30)-dependent ubiquitylation of Met4 controls Met4 activity remains contentious. Here, we have demonstrated that intracellular cysteine levels dictate the degradation of Met4 in vivo, as shown by the ability of cysteine, but not methionine or S-adenosylmethionine (AdoMet), to trigger Met4 degradation in an str4Delta strain, which lacks the ability to produce cysteine from methionine or AdoMet. Met4 degradation requires its nuclear localization and activity of the 26 S proteasome. Analysis of the regulated degradation of a fully functional Met4-Cbf1 chimera, in which Met4 is fused to the DNA binding domain of Cbf1, demonstrates that elimination of Met4 in vivo can be triggered independently of both its normal protein interactions. Strains that harbor the Met4-Cbf1 fusion as the only source of Cbf1 activity needed for proper kinetochore function exhibit high rates of methionine-dependent chromosomal instability. We suggest that SCF(Met30) activity or Met4 utilization as a substrate may be directly regulated by intracellular cysteine concentrations.
Menant
Alexandra
A
Centre de Génétique Moléculaire, Centre National de la Recherche Scientifique, 91198 Gif-sur-Yvettte, France.
Baudouin-Cornu
Peggy
P
Peyraud
Caroline
C
Tyers
Mike
M
Thomas
Dominique
D
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000235:genetics / Q000378:metabolism*; D050976:Basic-Leucine Zipper Transcription Factors / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D043171:Chromosomal Instability; D003545:Cysteine / Q000378:metabolism; D044783:F-Box Proteins; D015966:Gene Expression Regulation, Fungal; D018386:Kinetochores / Q000378:metabolism; D008715:Methionine / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D012436:S-Adenosylmethionine / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D013455:Sulfur / Q000378:metabolism*; 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; D050976:Basic-Leucine Zipper Transcription Factors; C064042:CBF1 protein, S cerevisiae; D044783:F-Box Proteins; C470285:MET30 protein, S cerevisiae; C073675:MET4 protein, S cerevisiae; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D013455:Sulfur; D012436:S-Adenosylmethionine; D008715:Methionine; D043743:Ubiquitin-Protein Ligase Complexes; D046988:Proteasome Endopeptidase Complex; D003545:Cysteine