Extended Data Fig. 4: Relationship between HSF1 foci and HSF1 transcriptional activity.
From: HSF1 phase transition mediates stress adaptation and cell fate decisions
a, Time-lapse heatmaps for HSF1-FI and HSP70 Transcriptional Reporter (TR) induction dTR/dt time derivative (one cell per row, n=152 cells). Black line, entry time of HSF1-FI < 0.2 b, Scatter plot of HSF1-FI versus dTR/dt from all instances from panel a (n=3,400 cell measurements, red line = linear fit). c,d, Scatter plot of TR versus (c.) time-integrated and (d.) maximum HSF1-FI 6 hours post 50nM STA9090 (Pearson correlation, bootstrapping p-values). e, Single-cell trace examples of time relationship between HSF1-FI and TR. Each dot represents a single-cell measurement, the color represents the time post STA9090 50nM addition (starting at 2 hours). Blue lines, linear fit y = α*x + β. f, Bootstrapping on parameter “α” in fits from panel e. Blue line, KD pdf of the α parameters from cells with R2>0.5. The matching between foci time dynamics and transcriptional reporter (TR) induction were randomly permuted 1,000 times. Red line, mean distribution of the α parameters for the permuted traces over all 1,000 permutations (light red area, 25th to 75th percentile). g, HSPA1A RNA FISH time course on U2OS HSF1-mEGFP cells post 100 nM STA9090 (n > 2200 cells per sample, see Source Data Extended Data Fig. 4 for statistics details). h, RNA FISH for HSPA1A on HCT15 HSF1-mVenus cells and U2OS HSF1-mEGFP cells after proteasome inhibition (Bortezomib 100 nM, 4 hours post). i-j, Immunofluorescence on U2OS HSF1-GFP cells (HSF1 green, HSP70 red, clone C92F3A-5). 1 µM MG132 overnight treatment. scalebar = 10 µm and j, quantification. Yellow, HSF1-FI<0.01, n = 1430 cells. Red, 0.01 < HSF1-FI < 0.2, n = 1156 cells. Blue HSF1-FI > 0.2, n = 492 cells. Inset, KD estimated distributions (two-sided KS p-values: high vs medium focied cells, p~10-41, medium vs low focied cells, p~10-18). Source data are provided in Source Data Extended Data Fig. 4.
