Journal: bioRxiv

Article Title: Regulation of ATAD2B bromodomain binding activity by the histone code

doi: 10.1101/2022.11.14.516501

Figure Lengend Snippet: (A) A surface representation of the ATAD2B BRD in complex with the H4K12ac ligand (residues 4-17) (PDB ID: 8EOQ). The H4K12ac ligand residues are shown as orange sticks, while the cartoon representation of BRD (cyan) highlights the ZA and BC loops in green and pink, respectively. (B) An isolated image of the simulated annealing composite omit map (gray) around the histone ligand H4K12ac (orange) contoured at 1σ. The composite omit map was calculated before building the histone ligand into the structure. All figures were made using the PyMOL Molecular Graphics system using version 2.3, Schrödinger, LLC. (C) Coordination of the H4K12ac ligand by the ATAD2B BRD. Residues lining the BRD pocket that are involved in ligand coordination are displayed in cyan, while the H4K12ac ligand residues are in orange. Hydrogen bond contacts are represented by a solid black line, and hydrophobic interactions are shown by dashed black lines. All contacts were determined using the PLIP program .

Article Snippet: CUT&RUN experiments were performed in the MCF7 cell line, using specific antibodies for the following proteins: ATAD2B (Invitrogen, Catalog # PA5-83233, Lot:), histone H4K5ac (Invitrogen, Ref # MA-32009; Lot:), histone H4K8ac (Epicypher Ab -Ref. 13-0036; Lot:), and histone H4K12ac (EpiCypher, SKU:13-0037, Lot:).

Techniques: Isolation

Journal: bioRxiv

Article Title: Regulation of ATAD2B bromodomain binding activity by the histone code

doi: 10.1101/2022.11.14.516501

Figure Lengend Snippet: (A) Immunoblot showing the interaction of GST-tagged ATAD2B BRD (41 kDa) with endogenous histone H4 (15 kDa) from MCF7 cells. Recombinant GST alone was used as a control. Specific antibodies were used for the GST tag, and the H4K5ac and H4K12ac modifications. (B) Euler plot showing strong replicate concordance for ATAD2B CUT&RUN samples. A total of 30,636 high-confidence peaks were identified across two biological replicates for ATAD2B. (C) Correlation heatmap based on comparing ATAD2B and our H4Kac PTM CUT&RUN data against all public ChIP-seq data for histone modifications in the breast cancer cell line MCF7. Clustering is derived from the Euclidian distance of signalValue for each factor and shows a strong correlation between ATAD2B and the H4Kac PTMs. (D) Signal heat map of CUT&RUN signals for all factors at the ATAD2B high-confidence peaks, showing strong signal correlation for histone H4Kac PTMs with the signal at ATAD2B peaks. (E) UCSC genome browser snapshots of CUT&RUN signal (reads per kilo-million) on multiple breast cancer-related genes (MYC, E2F1, BRCA1).

Article Snippet: CUT&RUN experiments were performed in the MCF7 cell line, using specific antibodies for the following proteins: ATAD2B (Invitrogen, Catalog # PA5-83233, Lot:), histone H4K5ac (Invitrogen, Ref # MA-32009; Lot:), histone H4K8ac (Epicypher Ab -Ref. 13-0036; Lot:), and histone H4K12ac (EpiCypher, SKU:13-0037, Lot:).

Techniques: Western Blot, Recombinant, Control, ChIP-sequencing, Derivative Assay