Extended Data Fig. 9: Condensation of DNAJB6b and FG-Nup fragments, and individual repetitions of the data in Fig. 6b.
a, DNAJB6b condensates by itself and is SDS soluble. Purified DNAJB6b was incubated with or without 10% PEG3350 (w/v). After 1 hour incubation, either 5% 2.5-hexanediol, 5% 1.6-hexanediol, or 0.5% SDS was added for 10 minutes and bright field images were taken. b, Fluorescence and bright field microscopy images showing co-localisation of Fluorescein-5-Maleimide labelled hNup153FG and yNup100FG in the absence or the presence of unlabelled DNAJB6b at a 3 μM concentration (molar ratio 1:1). Deconvolved fluorescence images are depicted as maximum Z-Projection of 30 consecutive slices of 0.2 micron. Scale bar represents 2 μm. c, Coomassie-stained SDS-PAGE showing one replicate of the sedimentation assay (quantified in Fig. 5e-f) to assess the soluble fraction of yNup100FG (3 μM) in the presence or absence of DNAJB6b (1.5 μM) for the indicated times. To determine the fraction 1,6-hexanediol soluble condensates, 10% 1,6-hexanediol was added for 10 minutes prior to centrifugation. Total yNup100FG protein is slightly shifted on the blot due to effects of PEG on mobility. d, Individual repetitions of the data in Fig. 6b. Filter-trap assays showing the effect of wildtype or indicated DNAJB6 mutants (18x S/T > A, 12x F > A, Δ79-115) on the aggregation of yNup100FG (3 μM; molar ratio 1:1). Graphs represent band intensities relative to the average intensity of the yNup100FG fragment alone (control). Source numerical data and unprocessed blots are available in source data.
