Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, United States; Center for Eukaryotic Gene Regulation, The Pennsylvania State University, University Park, United States; Microbiology Service, Department of Laboratory Medicine, National Institutes of Health Clinical Center, Bethesda, United States
Center for Eukaryotic Gene Regulation, The Pennsylvania State University, University Park, United States; Department of Chemistry, The Pennsylvania State University, Universtiy Park, United States
Yi Li
Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, United States; Center for Eukaryotic Gene Regulation, The Pennsylvania State University, University Park, United States
Samuel Becker
Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, United States
Fan Zou
Department of Physics, The Pennsylvania State University, University Park, United States
Xin Zhang
Department of Chemistry, The Pennsylvania State University, Universtiy Park, United States
Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, United States; Center for Eukaryotic Gene Regulation, The Pennsylvania State University, University Park, United States; Department of Physics, The Pennsylvania State University, University Park, United States
Some transcription factors (TFs) can form liquid–liquid phase separated (LLPS) condensates. However, the functions of these TF condensates in 3-Dimentional (3D) genome organization and gene regulation remain elusive. In response to methionine (met) starvation, budding yeast TF Met4 and a few co-activators, including Met32, induce a set of genes involved in met biosynthesis. Here, we show that the endogenous Met4 and Met32 form co-localized puncta-like structures in yeast nuclei upon met depletion. Recombinant Met4 and Met32 form mixed droplets with LLPS properties in vitro. In relation to chromatin, Met4 puncta co-localize with target genes, and at least a subset of these target genes is clustered in 3D in a Met4-dependent manner. A MET3pr-GFP reporter inserted near several native Met4-binding sites becomes co-localized with Met4 puncta and displays enhanced transcriptional activity. A Met4 variant with a partial truncation of an intrinsically disordered region (IDR) shows less puncta formation, and this mutant selectively reduces the reporter activity near Met4-binding sites to the basal level. Overall, these results support a model where Met4 and co-activators form condensates to bring multiple target genes into a vicinity with higher local TF concentrations, which facilitates a strong response to methionine depletion.
