Efficient Pre-mRNA Cleavage Prevents Replication-Stress-Associated Genome Instability

Mol Cell. 2019 Feb 21;73(4):670-683.e12. doi: 10.1016/j.molcel.2018.11.036. Epub 2019 Jan 10.

Abstract

Cellular mechanisms that safeguard genome integrity are often subverted in cancer. To identify cancer-related genome caretakers, we employed a convergent multi-screening strategy coupled to quantitative image-based cytometry and ranked candidate genes according to multivariate readouts reflecting viability, proliferative capacity, replisome integrity, and DNA damage signaling. This unveiled regulators of replication stress resilience, including components of the pre-mRNA cleavage and polyadenylation complex. We show that deregulation of pre-mRNA cleavage impairs replication fork speed and leads to excessive origin activity, rendering cells highly dependent on ATR function. While excessive formation of RNA:DNA hybrids under these conditions was tightly associated with replication-stress-induced DNA damage, inhibition of transcription rescued fork speed, origin activation, and alleviated replication catastrophe. Uncoupling of pre-mRNA cleavage from co-transcriptional processing and export also protected cells from replication-stress-associated DNA damage, suggesting that pre-mRNA cleavage provides a mechanism to efficiently release nascent transcripts and thereby prevent gene gating-associated genomic instability.

Keywords: ATR; R-loops; RNA:DNA hybrids; checkpoint activation; cleavage; gene gating; origin firing; polyadenylation; pre-mRNA processing; replication catastrophe; replication stress.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Active Transport, Cell Nucleus
  • Cell Cycle Proteins / genetics
  • Cell Cycle Proteins / metabolism
  • DNA Damage*
  • DNA Replication*
  • DNA, Neoplasm / genetics
  • DNA, Neoplasm / metabolism
  • Gene Expression Regulation, Neoplastic
  • Genomic Instability*
  • HeLa Cells
  • Humans
  • Neoplasms / genetics*
  • Neoplasms / metabolism
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism
  • Nucleic Acid Heteroduplexes / genetics
  • Nucleic Acid Heteroduplexes / metabolism
  • Polyadenylation
  • RNA Cleavage*
  • RNA Precursors / biosynthesis
  • RNA Precursors / genetics*
  • RNA, Messenger / biosynthesis
  • RNA, Messenger / genetics*
  • RNA, Neoplasm / biosynthesis
  • RNA, Neoplasm / genetics*

Substances

  • Cell Cycle Proteins
  • DNA, Neoplasm
  • Nuclear Proteins
  • Nucleic Acid Heteroduplexes
  • RNA Precursors
  • RNA, Messenger
  • RNA, Neoplasm
  • THOC1 protein, human
  • WDR33 protein, human