Fig. 4: HMGA2 forms droplets with nucleosomes.

a The disorder score was calculated with IUPred3 or ANCHOR2^92,93. b FRAP analysis of HMGA2-EGFP in a neocortical cell prepared from the Hmga2-EGFP mouse. The EGFP signal at the bleached focus (indicated by the dotted circle) was mostly recovered by 40 s after bleaching (postbleach). Scale bar, 5 µm. c Quantification of EGFP signal intensity at the bleached focus in FRAP experiments as in (b) (photobleaching ended at t = 0). Data are means ± s.d. (n = 72 droplets from 72 cells, n = 3 independent experiments). d Droplet formation analysis. Recombinant HMGA2 (20 μM) and mononucleosomes (800 nM) were incubated alone or together in a droplet formation solution and then observed with a differential interference contrast (DIC) microscope. Scale bars, 10 µm. e Turbidity of incubation mixtures as in (d) was assessed by measurement of optical density at 400 nm (OD₄₀₀). Data are means ± s.d. (n = 3 independent experiments). f Phase diagram of HMGA2 droplet formation. HGMA2 at the indicated concentration was added to a droplet formation buffer containing the indicated concentration of NaCl and OD₄₀₀ was measured after 10 min (n = 3 independent experiments). g, h Droplet formation by and FRAP analysis of HMGA2 labeled with a fluorescent tag. Fluorescence microscopy of droplets formed by 20 μM ATTO647-labeled recombinant HMGA2 (labeling efficiency of <30%) and 800 nM mononucleosomes is shown in (g). Scale bar, 10 µm. Such droplets were subjected to FRAP analysis (postbleach corresponds to 55 s after bleaching), with the signal intensity having recovered to 80.2% ± 2.0% (mean ± s.d., n = 9 from three independent experiments) by 30 s after bleaching (h). Scale bar, 1 µm.