Extended Data Fig. 10: Swi6 dimerX and loopX mutants are defective in promoting octamer distortion and forming heterochromatin foci in S. pombe.
From: HP1 reshapes nucleosome core to promote phase separation of heterochromatin
a, Nucleosome binding assays by fluorescence anisotropy showing that binding of Swi6 loopX mutant to H3K9cme3 nucleosomes is not affected by H2B–H4 histone disulfide linkage. H2B•H4 S-S are oxidized nucleosome, H2B•H4 S-H are reduced nucleosomes. Kd values are shown. b, Nucleosome binding assays by fluorescence anisotropy showing that binding of the Swi6 loopX and dimerX mutants to H3K9me3 nucleosomes is not affected by H3–H4 histone disulfide linkage. H3•H4 S-S are oxidized nucleosome, H3•H4 S-H are reduced nucleosomes. Kd values are shown. c, Wild-type Swi6 and loopX and dimerX mutant Swi6 proteins do not form liquid droplets in absence of chromatin (Swi6 concentration is 4 μM). d, Bright-field images showing droplet wetting over time. Swi6–chromatin condensates are incubated at room temperature and images are collected at the indicated times. Condensates are imaged on a bottom glass plate coated with Peg-silane (see Methods). e, Swi6 immunofluorescence on dcr1Δ swi6Δ S. pombe cells shows absence of Swi6 staining, confirming the specificity of the anti-Swi6 antibody. Images in c–e are representative of at least three independent experiments. In a and b, measurements entailed three independent experiments; error bars denote s.d.
