ATR inhibition rewires cellular signaling networks induced by replication stress

Area of Science:

• Molecular Biology
• Cellular Biology
• Cancer Research

Background:

• Replication stress, including fork stalling, stems from DNA lesions, nucleotide depletion, and oncogene activation.
• The ataxia telangiectasia and Rad3-related kinase (ATR) is crucial for the cellular response to replication stress.
• ATR inhibition is a promising cancer therapeutic strategy for tumors with high replication stress.

Purpose of the Study:

• To systematically investigate ATR's substrate spectrum and the signaling networks induced by replication stress.
• To define cellular signaling following nucleotide depletion-induced replication stress and ATR inhibition.
• To identify novel ATR targets and understand the rewiring of signaling networks upon ATR inhibition.

Main Methods:

• Quantitative mass spectrometry-based proteomics was used to analyze cellular signaling.
• Proteomic analysis was performed after inducing replication stress (nucleotide depletion) and inhibiting ATR.
• Identification of phosphorylated proteins and putative ATR phosphorylation sites.

Main Results:

• Replication stress increases phosphorylation of proteins involved in RNA splicing and transcription, previously unlinked to replication stress response.
• ATR-dependent phosphorylation events were identified following replication stress.
• Novel ATR target sites were discovered on MCM6, TOPBP1, RAD51AP1, and PSMD4.
• ATR inhibition was shown to rewire replication stress-induced signaling networks, activating ATM-driven double-strand break repair.

Conclusions:

• Replication stress triggers novel signaling pathways involving RNA splicing and transcription.
• ATR plays a key role in phosphorylating specific proteins during replication stress.
• ATR inhibition alters cellular signaling, leading to ATM pathway activation, offering insights into cancer treatment strategies.