Fig. 5: S47 O-GlcNAcylation orients MCM association with H4 on chromatin.
From: H4S47 O-GlcNAcylation regulates the activation of mammalian replication origins
a,b, In vitro assay of MCM2 interaction with H4 and DDK. Purified GST-MCM2 (a) and His-CDC7 and His-DBF4 (b) were incubated with His-H4WT or His-H4S47A, which were isolated from BL21 competent E. coli cells co-expressing OGT. c, Co-IP analysis for H4 association with MCM2, MCM7, and DBF4. HEK293T cells, prepared as in Figure 1a, were pre-extracted by CSK-0.5% Triton and immunoprecipitated with Flag antibody. d, Assay of H4-MCM2 interaction by PLA. HEK293T cells were prepared as in Figure 1a and pre-extracted with CSK-0.5% Triton prior to PLA. CAF1p150-PCNA and IgG-Flag interactions served as positive and negative controls, respectively. Representative images (left) and dot plots showing intensities of nuclear PLA foci (right) are shown. From left, n = 3,038, 1,902, 1,866, 2,000, 1,858, 1,699 cells. P values were calculated by unpaired, two-tailed t-tests, means ± s.d.; red bars indicate mean values. Scale bar, 5 μm. a.u., arbitrary unit.
