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This deposition contains supplementary data files and laboratory notebooks of the manuscript 'Systematic dissection of the regulatory logic of transcriptional activation by TP53 binding sites'. In this study, thousands of barcoded TP53 reporters were systematically designed and probed in MCF7 and TP53-KO MCF7 cells upon stimulation with Nutlin-3a. Barcodes in the cDNA were quantified by high-throughput sequencing. Among others, the results demonstrate that TP53 drives transcription cooperatively from adjacent TP53 binding sites, and that the positioning of the TP53 binding sites is crucial for transcriptional activation. The files deposited here include: Laboratory notebooks (pdfs printedfrom labguru). This includes the generation of the plasmid pools, the library transfection protocols, and the western blotting procedure. Raw gel images etc. are available in this folder. Clustered barcode count files. These are the raw barcode counts that were extracted from the sequencing files and clustered using starcode. A reporter information sheet. This file includes a table containing all relevant reporter information and reporter sequences. A reporter activity sheet. This file includes the calculated reporter activities and can be used to recreate the figures presented in the manuscript. A supplementary table containing the primer sequences used in this study. &nbsp

The efficiency and outcome of CRISPR/Cas9 editing depends on the chromatin state at the cut site. It has been shown that changing the chromatin state can influence both the efficiency and repair outcome, and epigenetic drugs have been used to improve Cas9 editing. However, because the target proteins of these drugs are not homogeneously distributed across the genome, the efficacy of these drugs may be expected to vary from locus to locus. Here, we systematically analyzed this chromatin context-dependency for 160 epigenetic drugs. We used a human cell line with 19 stably integrated reporters to induce a double-stranded break (DSB) in different chromatin environments. We then measure Cas9 editing efficiency and repair pathway usage by sequencing the mutational signatures. We identified 67 drugs that modulate Cas9 editing efficiency and/or repair outcome dependent on the local chromatin environment. For example, we find a subset of histone deacetylase inhibitors that improve Cas9 editing efficiency throughout all types of heterochromatin (e.g., PCI-24781), while others were only effective in H3K27me3-marked regions (e.g., Vorinostat). In summary, this study reveals that most epigenetic drugs alter CRISPR editing in a chromatin-dependent manner, and provides a detailed guide to improve Cas9 editing more selectively at the desired location.HIGHLIGHTSA screen identifies dozens of drugs that alter Cas9 editing in a chromatin context-dependent mannerMany HDAC inhibitors boost Cas9 editing efficiency throughout all types of heterochromatinThe DNMT inhibitor Decitabine completely blocks resection-dependent repair across the genome

This deposition contains supplementary data files and laboratory notebooks of the manuscript 'Systematic dissection of the regulatory logic of transcriptional activation by TP53 binding sites'. In this study, thousands of barcoded TP53 reporters were systematically designed and probed in MCF7 and TP53-KO MCF7 cells upon stimulation with Nutlin-3a. Barcodes in the cDNA were quantified by high-throughput sequencing. Among others, the results demonstrate that TP53 drives transcription cooperatively from adjacent TP53 binding sites, and that the positioning of the TP53 binding sites is crucial for transcriptional activation. The files deposited here include: Laboratory notebooks (pdfs printedfrom labguru). This includes the generation of the plasmid pools, the library transfection protocols, and the western blotting procedure. Raw gel images etc. are available in this folder. Clustered barcode count files. These are the raw barcode counts that were extracted from the sequencing files and clustered using starcode. A reporter information sheet. This file includes a table containing all relevant reporter information and reporter sequences. A reporter activity sheet. This file includes the calculated reporter activities and can be used to recreate the figures presented in the manuscript. A supplementary table containing the primer sequences used in this study. &nbsp

Contains all uniquely mapped TRIP barcode integration sites used in Leemans et al. with their LAD-state and expression

Contains annotation of all enhancers used in this experiment with SuRE expression and classification.

Tabular data files with SuRE counts, for 4 SuRE libraries, 2 bio reps each

Contains annotation of all promoters used in this experiment with SuRE expression and classification.

4 text files with lists of SNPid's of significant SNPs and matched controls, for K562 and HepG2 cell lines

Coordinates for LAD atlas in tabular txt files