Fig. 1: Recombinant TFG assembles into anisotropic, hollow condensates in vitro.
From: A hollow TFG condensate spatially compartmentalizes the early secretory pathway
a Representative 4-color micrograph of an individual ERES/cis-Golgi unit. HeLa cells were transfected with mGFP-Sec16L (green), FLAG-TFG-SNAP (cyan) (labeled with SNAP-Cell 647-SiR), and immunostained for endogenous GM130 (gray) and TANGO1 (red). Scale bar 1 µm. b Fluorescence intensity line scan of 4-color ERES/cis-Golgi unit (averaged and normalized to TANGO1, n = 4). Schematic of localizations detected in the representative unit (a) is given. c Representative purity of peak TFG elutions from concentration/salt exchange (Coomassie staining). d Confocal time series of increasing concentration of TFG-mEGFP-FLAG at the edge of an evaporating 10 µL sessile droplet, 150 mM KCl. Scale bar 5 μm. Starting concentration ~100 nM. e TFG-mEGFP-FLAG at 430 nM spontaneously assembles into ‘sponge-like’, anisotropic condensates (HEPES/KOH pH 7.3; 150 mM KCl; 20% (v/v) PEG 8 kDa; confocal microscopy). Scale bar 5 µm. Individual TFG condensate, Z-step 125 nm. Scale bar 500 nm. f Schematic depicting the ER–Golgi interface (targets for immunolabeling at ERES: red; cis-Golgi: grey. g ERES to cis-Golgi distance measured between immunolabeled GM130 and TANGO1, mean = 269, n = 120, vs diameter of individual hollow TFG condensates in vitro, mean = 341, n = 175. h Determination of the saturation concentration required for the formation of hollow TFG condensates (in the absence of crowding agents). Representative micrographs depicting the distribution of TFG below and above the saturation concentration of 100 nM. Higher concentrations of TFG yield inverted phases (right micrograph). Dilute phase (left) shown at <5 nM, dense phase (middle) shown at 200 nM, inverted phase (right) shown at 6.4 µM. Red diamonds indicate saturation concentration and average cellular concentration of TFG. Scale bars 5 µm. Source data are provided as a Source Data file.
