Extended Data Fig. 2: PDHE1α-catalyzed local acetyl-CoA is required for chromatin acetylation in response to DNA damage.
From: PARylated PDHE1α generates acetyl-CoA for local chromatin acetylation and DNA damage repair
a, Dynamics of citrate metabolism at DNA damage sites monitored by laser micro-irradiation-coupled live-cell imaging on HeLa cells overexpressing the highly responsive citrate sensor construct, NLS-Citron1/pCDNA3.1. The fluorescence intensity at DSB stripes was quantificated from three independent replicates (n = 20 for each group), the curve shows the mean ± s.e.m. Scale bars, 5 μm. b-c, Volcano plots of the acetyl-modified proteins identified in Fig. 2B. Relative acetylation levels in the IR (b) and 4-OHT (c)-induced DNA damage groups versus the control are plotted on the x-axis as mean log2 ratios. Negative log10 PEP (the maximal posterior error probability for peptides) values are plotted on the y-axis. Significantly enriched proteins are denoted by red and blue dots, 13C isotope-labeled proteins are denoted by green dots, and all others are denoted by gray dots. d-e, Venn diagram showing all identified (d) and 13C isotope-labeled (e) that overlap between significant lysine acetylation sites (upper) and between acetylated proteins (lower) in IR- and 4-OHT-induced DNA damage versus control groups. f, Western blot showing the histone acetylation marks in acid-extracted fractions and the knockdown efficiency in whole cell lysate from HeLa cells transfected with siRNAs targeting PDHE1α, ACLY, and ACSS2 or NC. g-h, Immunofluorescence staining showing the colocalization of γH2AX with pan-Ac (g) and H3K9ac (h) signal in laser micro-irradiatiation induced DNA damage sites, nuclei were counterstained with DAPI. Pan-lysine acetylation intensity at DSB stripes was quantified from three independent replicates where data were collected from 78 cells for g and 51 cells for h group. The statistical analyses were performed using a two-tailed student’s t-test, and shows the mean ± s.d. Scale bars, 5 μm. i, Quantifition of total 13C-isotope -abelled Kac intensity around DSB sites in 13C-pyruvate, 13C-acetate and 13C-citrate cultured cells.
