Fig. 3.

Double N-tail delete, tH2A:tH3 mutant cells have more open chromatin structure. (A) Bulk chromatin from wild-type and tH2A:tH3 cells was digested with different amounts of MNase (lanes 1 and 5, 0 U; lanes 2 and 6, 1 U; lanes 3 and 7, 2 U; lanes 4 and 8, 4 U) and visualized with ethidium bromide on agarose gel. M, DNA ladders; U, relative units of Mnase. (B) Densitometric analysis of MNase-digested DNA shown in A was measured by plotting profile with Image J (National Center for Biotechnology Information, NCBI). Dashed lines mark mono- (n1), di- (n2), and tri- (n3) nucleosomes. Labels for plotted profile (1–8) are matched with lanes in A. (C) Chromatin assembly, as assayed by superhelical density analysis of 2-μm plasmids (ectopically introduced pRS426), is shown in histone-tail deletion mutants and wild-type cells. Samples from histone shuffle strain-derived cells were run from lanes 1–6 (lane 1, wild type; lane 2, tH2A; lane 3, tH2B; lane 4, tH3; lane 5, tH2A:tH3; lane 6, tH2B:tH3). Lanes 7 and 8 contain samples from BY4741 (wild type with two endogenous copies of each wild-type core histones) and cac1Δ derived from BY4741, respectively. N, nicked plasmids and the distributed topoisomers of the plasmid are marked with a bracket. The midpoint of topoisomer distribution in each lane (marked with a closed circle) was found by profiling the toposiomers with Image J (NCBI).