biostudies-literatureLee JNational Institute of General Medical SciencesNIGMS NIH HHSNational Science FoundationRP96028https://www.ebi.ac.uk/biostudies/studies/S-EPMC11441978biostudies-literatureUnknown13Some 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 <i>MET3pr</i>-GFP reporter inserted near several native MeLifeTranscription factor condensates, 3D clustering, and gene expression enhancement of the <i>MET</i> regulon.PMC11441978R35 GM139654MCB 2016266Li YZhang XLee JBai LBecker SZou FSimpson LfalseTranscription factor condensates, 3D clustering, and gene expression enhancement of the <i>MET</i> regulon.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 <i>MET3pr</i>-GFP reporter inserted near several native M2024-01-01T00:00:00Z2024 Sep2026-07-15T13:10:36.498Z2025-04-06T12:29:29.953ZS-EPMC114419783934773810.7554/eLife.96028