Fig. 2. Celastrol promotes phase separation and liquidity of p62.

a Representative images showing the time-dependent effect on celastrol induction of cytoplasmic Nur77 body formation. Bottom panels: quantitative analysis of the number and size of Nur77/p62 body formation. Bottom left graph, n = 4 biologically independent samples; Bottom right graph, n = 20, 23, 25, and 19, respectively. Data were presented as mean values ± SEM. Scale bar, 10 μm. b Real-time images showing the formation and fusion of GFP-Nur77 and mCherry-p62 droplets in HeLa cells after treatment with celastrol (2 μM) for 1 h. White arrows indicate droplets formation and fusion (see also Supplementary Movie 1). Scale bar, 10 μm. c Representative images illustrating the role of celastrol in promoting p62 body formation in a Nur77-dependent manner immunostaining. Nur77^−/−HeLa cells were also transfected with GFP-Nur77 to determine its effect on p62 body formation. The diameter of the biggest p62 puncta in each cell was measured. The number of p62 puncta >0.5 μm in each cell was assessed. A two-tailed unpaired Student’s t-test was used for statistical analysis, and data are presented as mean values ± SEM (n = 5 biologically independent samples). d FRAP analysis of the effect of Nur77 in regulating p62 mobility in HeLa cells. Data were presented as means ± SEM (n = 3 independent experiments). Scale bar, 1.5 μm. Data represent at least three independent experiments. Source data are provided as a Source Data file.