Fig. 4: Hydrophobic ‘sticker’ residues control the anisotropic condensation mechanism of TFG.

a Schematic depicting domains present within TFG: PB1 domain, blue; alpha helices, green; Gln-rich region, purple; intrinsically disordered region (IDR), gray. Individual IDRs are defined as: IDR1 (net acidic), IDR2 (Gln-rich), IDR3 (net basic). Residues for truncated constructs are depicted to scale. b All truncated constructs except for residues 1-123 of TFG support the formation of lumen-containing (anisotropic) condensates. Representative micrographs are given. Overview scale bars 5 µm; insets 500 nm. c Quantification of the average peak-to-peak diameter of condensates formed by individual truncations of TFG. TFG; TFG 124–240: n = 124; TFG 124–161: n = 79; TFG 162–240: n = 176; TFG 241–359: n = 93; TFG 360–400: n = 107. The dashed line represents the average TFG 1–400 condensate diameter for reference. Two-tailed unpaired t-tests (TFG-124–240: p < 0.0001, TFG-185–240: p = 0.0006, TFG-241–359: p = 0.0011, TFG-360–400: p < 0.0001). d TFG phase diagram probing increasing concentrations of KCl vs PEG 8 kDa as a crowding agent. TFG-mEGFP-FLAG: 70 nM; HEPES/KOH pH 7.3; X mM KCl as indicated; X% (v/v) PEG 8 kDa as indicated. Scale bar 10 µm. e Schematic based on an AlphaFold prediction of TFG structure color-coded for hydrophobicity (red: max; blue: min). Domain annotations and the position of hydrophobic residues (‘stickers’) within IDRs are given. f Widefield deconvolution micrographs of HeLa cells transfected with TFG-mEGFP-FLAG and TFG-1–400-w/o ‘stickers’-mEGFP-FLAG, respectively. Scale bar 10 µm. g Representative micrographs of HeLa cells transfected with TFG-mEGFP-FLAG shifted to the indicated temperatures (top) and time-lapse of a single cell shifted through indicated temperatures (bottom). Scale bar 10 µm. h Quantification of condensate circularity at specific temperatures. Two-tailed unpaired t-tests (^*p = 0.0378, ^***p = 0.0003, ^****p < 0.0001) i Micrographs of TFG-162-240-mEGFP-FLAG in vitro incubated at indicated temperatures. Representative of three experiments. Scale bar 5 µm. Source data are provided as a Source Data file.