Fig. 4: The molecular mechanisms of p300BRPHZ phase separation. | Nature Communications

Fig. 4: The molecular mechanisms of p300BRPHZ phase separation.

From: Nuclear condensates of p300 formed though the structured catalytic core can act as a storage pool of p300 with reduced HAT activity

Fig. 4

a Representative images of untreated or SIRT2-deacetylated N1132A p300BRPHZ samples containing 13 µM protein and 12% PEG in a 50 µm × 50 µm square region on cover slides under the microscope. b Quantification of droplets counted in images acquired for the sample in (a). The number of droplets was counted in five non-overlapping 50 µm × 50 µm square regions and the mean value was plotted. Error bar represents SD. c Schematic of the N1132A p300BRPHZ phase separation via the intermolecular interaction between the HAT domain and deacetylated AIL. d Representative images of untreated or SIRT2-deacetylated ΔAIL p300BRPHZ samples containing 13 µM protein and 12% PEG in a 50 µm × 50 µm square region on cover slides under a microscope. e Quantification of droplets counted in images acquired for the sample in (d). The number of droplets was counted in five non-overlapping 50 µm × 50 µm square regions and the mean value was plotted. Error bar represents SD. f Schematic of the ΔAIL p300BRPHZ phase separation via the intermolecular interaction between BD and acetylated lysines outside AIL. g A representative image of purified D1399A p300BRPHZ sample (from at least three replicates) containing 13 µM protein and 12% PEG in a 50 µm × 50 µm square region on cover slides under the microscope. h Schematic of the catalytically inactive D1399A p300BRPHZ phase separation via the intermolecular interaction between the HAT domain and unmodified AIL. i Representative images of untreated or SIRT2-deacetylated WT p300HZ samples containing 13 µM protein and 12% PEG in a 50 µm × 50 µm square region on cover slides under the microscope. j Schematic of the p300HZ phase separation via the intermolecular interaction between the HAT domain and deacetylated AIL. k Representative images of untreated or SIRT2-deacetylated ΔAIL p300HZ samples containing 13 µM protein and 12% PEG in a 50 µm × 50 µm square region on cover slides under the microscope. l Quantification of droplets counted in images acquired for the samples in (i, k). The number of droplets was counted in five non-overlapping 50 µm × 50 µm square regions and the mean value was plotted. Data are presented as mean values ± SD; the error bar represents SD. m Representative images of HeLa cells (from at least three replicates) expressing N1132A and ΔAIL mutants of YFP-p300FL. Scale bar, 5 µm.

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