This notebook contains high level information on the bioinformatics pipeline that was used for the analysis reported in Szydzik J et al. ATR inhibition enables complete tumour regression in ALK-driven NB mouse models, Nature Communications 2021

Environment

Analysis was performed in a Conda environment. See ATR.yml for details. scripts/Rpacks describes R packages that were installed independently.

Experimental conditions

RNA-Seq data cell lines after ATR inhibition with 50nM BAY 1895344

• CLB-BAR: 0h (ctrl), 24h, 48h
• CLB-GE: 0h (ctrl), 24h, 48h

Proteomics data cell lines after ATR inhibition with 50nM BAY 1895344

• CLB-GE: 0h (ctrl), 24h, 48h
• CLB-BAR: 0h (ctrl), 24h, 48h

Phosphoproteomics data cell lines after ATR inhibition with 50nM BAY 1895344

• CLB-BAR: SYNCHRONIZED, 6h (DMSO), 6h (Treatment)

RNA-Seq data ALK and ALKAL mice after 3d ATR inhibition with 50 nM BAY 1895344.

Data

• Raw RNA-Seq data have been deposited in Arrayexpress
  • Cell lines: E-MTAB-10603
  • Mice: E-MTAB-10616 (BAY treatment) & E-MTAB-9600 (Control samples)
• Processed data are available in data/ATR_data.RData. This file containes the following objects:
  • normalized_counts: DESeq2-normalized counts from cell line RNA-Seq data
  • res_diff_expr: DESeq2 output from cell line RNA-Seq data
  • normalized_counts_prot: Normalized counts from cell line proteomics data
  • res_diff_expr_prot: DEP output from cell line proteomics data
  • normalized_counts_PP: Normalized counts from cell line phosphoproteomics data
  • res_diff_expr_PP: DEP output from cell line phosphoproteomics data
  • sample_info: sample information cell line RNA-Seq data
  • sample_info_prot: sample information cell line (phospho-)proteomics data
  • normalized_counts_mice: DESeq2-normalized counts from mice RNA-Seq data
  • res_diff_expr_mice: DESeq2 output from mice RNA-Seq data
  • sample_info_mice: sample information mice RNA-Seq data
  • SQ_sites: Information on S/TQ sites of phosphoproteomics data
  • geneset_ls: genesets downloaded from MSigDB v7.2
  • MGI_to_HGNC: MGI to HGNC mapping table
  • ppi_db: High confidence interactions from STRING

Manuscript

The main analysis, as reported in the manuscript

RNA-Seq cell lines

source("scripts/manuscript_RNA.R")

Proteomics cell lines

source("scripts/manuscript_prot.R")

PP cell lines

Basic analysis: volcano plot, ST/Q sites, GSEA (+ table)

source("scripts/manuscript_PP.R")

Heatmap with main results

source("scripts/manuscript_PP_hm.R")

Network analysis

source("scripts/manuscript_PP_network.R")

Motif analysis

source("scripts/manuscript_PP_motifs.R")

RNA-Seq mice

source("scripts/manuscript_mice.R")

Summary table

source("scripts/manuscript_summary.R")