Fig. 3: MeCP2 preferentially and stably associates with nucleosomes on chromatinized DNA.
From: Differential dynamics specify MeCP2 function at nucleosomes and methylated DNA
a, Schematic of the experimental setup. LD655-labeled nucleosomes and Cy3-labeled MeCP2 were simultaneously monitored via two-color confocal fluorescence microscopy. b,c, Representative kymograph of a nucleosome-containing unmethylated (b) or CpG methylated (c) DNA tether incubated with 2 nM Cy3-MeCP2. A red laser was flashed on briefly to locate the nucleosomes within the tether. The arrowheads denote nucleosome positions. The experiment was independently repeated with 22 (unmethylated) and 18 (methylated) tethers yielding similar results. d, Average MSD plot for Cy3-MeCP2 trajectories on unmethylated bare DNA (n = 78 from 22 independent tethers), methylated bare DNA (n = 200 from 18 independent tethers) or colocalized with nucleosome loci on unmethylated chromatinized DNA (n = 22 from 6 independent tethers). The error bars represent s.d. e, Estimated number of Cy3-MeCP2 molecules per trajectory on unmethylated bare DNA or bound to nucleosomes on unmethylated DNA. The bars represent mean and s.d. Significance was calculated using a two-tailed unpaired t-test. f, Fraction of Cy3-MeCP2 trajectories that were colocalized with nucleosome loci on unmethylated DNA in the presence of 0.5 nM (from 5 independent tethers), 1 nM (from 6 independent tethers), 2 nM (from 9 independent tethers) or 6 nM (from 8 independent tethers) MeCP2. The bars represent mean and s.d. Significance was calculated using a two-tailed unpaired t-test for each pair. g,h, Representative kymograph of a nucleosome-containing unmethylated (g) or CpG methylated (h) DNA tether incubated with 6 nM Cy3-MeCP2. The arrowheads denote nucleosome positions. The experiment was independently repeated with 8 (unmethylated) and 7 (methylated) tethers yielding similar results.
