Transcription-replication conflicts as a source of common fragile site instability caused by BMI1-RNF2 deficiency.

Anthony Sanchez,Peter Tonzi,Angelo de Vivo,Younghoon Kee,Jeonghyeon Kim,Tony T Huang,Charis Eng

doi:10.1371/journal.pgen.1008524

Anthony Sanchez, Peter Tonzi + Show 5 more

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https://doi.org/10.1371/journal.pgen.1008524

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Journal: PLoS Genetics	Publication Date: Mar 6, 2020
Citations: 21	License type: CC BY 4.0

Affiliation: University of South Florida, New York University

Abstract

Common fragile sites (CFSs) are breakage-prone genomic loci, and are considered to be hotspots for genomic rearrangements frequently observed in cancers. Understanding the underlying mechanisms for CFS instability will lead to better insight on cancer etiology. Here we show that Polycomb group proteins BMI1 and RNF2 are suppressors of transcription-replication conflicts (TRCs) and CFS instability. Cells depleted of BMI1 or RNF2 showed slower replication forks and elevated fork stalling. These phenotypes are associated with increase occupancy of RNA Pol II (RNAPII) at CFSs, suggesting that the BMI1-RNF2 complex regulate RNAPII elongation at these fragile regions. Using proximity ligase assays, we showed that depleting BMI1 or RNF2 causes increased associations between RNAPII with EdU-labeled nascent forks and replisomes, suggesting increased TRC incidences. Increased occupancy of a fork protective factor FANCD2 and R-loop resolvase RNH1 at CFSs are observed in RNF2 CRISPR-KO cells, which are consistent with increased transcription-associated replication stress in RNF2-deficient cells. Depleting FANCD2 or FANCI proteins further increased genomic instability and cell death of the RNF2-deficient cells, suggesting that in the absence of RNF2, cells depend on these fork-protective factors for survival. These data suggest that the Polycomb proteins have non-canonical roles in suppressing TRC and preserving genomic integrity.

Highlights

Common fragile sites (CFSs) are natural genomic loci that are prone to gaps and breaks upon DNA replication stress
We found that deficiency in the Polycomb group proteins BMI1 or RNF2 leads to the CFS instability, and is associated with transcriptionassociated replication fork stresses
BMI1-RNF2 suppress transcription-induced instability at common fragile sites depend on the Fanconi Anemia fork-protective proteins for genome maintenance and survival

Summary

Introduction

Common fragile sites (CFSs) are natural genomic loci that are prone to gaps and breaks upon DNA replication stress. A few potential sources for the replication defects and increased breakages at CFSs have been proposed, such as high frequencies of DNA secondary structures forming barriers to the fork progression, scarcity of replication origins, and collisions between transcription and replication [2,3,4,5]. Prokaryotic and eukaryotic cells appear to have evolved mechanisms to prevent transcription-replication conflicts (TRCs), by separating the timing and location of transcription or replication processes [10]. This may be challenging at long genes where the transcription of a single long gene can take place throughout the entire cell cycle including when replication is active [8]. TRC incidences can be accelerated by the overexpression of oncogenes such as MYC [11], RAS [12], or Cyclin E [13], which could alter replication origin firing or global transcription

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