Dynamics of replication origin over-activation

Haiqing Fu,Bhushan L Thakur,Robin Sebastian,Mishal Rao,Shira T Mencer,Sophie Z Zhuang,Sang-Min Jang,Mirit I Aladjem,Sarah B Lazar,Christophe E Redon,Lorinc S Pongor,Anna B Marks,Jacob M Gross,Sara Mosavarpour,Koichi Utani,Adrian M Baris

doi:10.1038/s41467-021-23835-0

Abstract

Safeguards against excess DNA replication are often dysregulated in cancer, and driving cancer cells towards over-replication is a promising therapeutic strategy. We determined DNA synthesis patterns in cancer cells undergoing partial genome re-replication due to perturbed regulatory interactions (re-replicating cells). These cells exhibited slow replication, increased frequency of replication initiation events, and a skewed initiation pattern that preferentially reactivated early-replicating origins. Unlike in cells exposed to replication stress, which activated a novel group of hitherto unutilized (dormant) replication origins, the preferred re-replicating origins arose from the same pool of potential origins as those activated during normal growth. Mechanistically, the skewed initiation pattern reflected a disproportionate distribution of pre-replication complexes on distinct regions of licensed chromatin prior to replication. This distinct pattern suggests that circumventing the strong inhibitory interactions that normally prevent excess DNA synthesis can occur via at least two pathways, each activating a distinct set of replication origins.

Highlights

Safeguards against excess DNA replication are often dysregulated in cancer, and driving cancer cells towards over-replication is a promising therapeutic strategy
We have previously shown that re-replication could be triggered by inhibiting Skp[2], a key component of CRL1, in a RepID-deficient cell background that prevented the recruitment of the alternative ubiquitin ligase CRL4 to chromatin[23]
Unlike normal mitotically growing cells, which activate a fraction of their potential replication origins on each chromosome each cell division, resulting in inter-origin spacing of 100–150 kb, we found that re-replicating cells show a higher frequency of initiation from origins positioned in closer proximity to one another

Summary

Introduction

Safeguards against excess DNA replication are often dysregulated in cancer, and driving cancer cells towards over-replication is a promising therapeutic strategy. The skewed initiation pattern reflected a disproportionate distribution of pre-replication complexes on distinct regions of licensed chromatin prior to replication This distinct pattern suggests that circumventing the strong inhibitory interactions that normally prevent excess DNA synthesis can occur via at least two pathways, each activating a distinct set of replication origins. Excess potential origins that are bound by pre-replication complexes do not initiate replication during normal mitotic growth (dormant origins) These dormant origins can be activated under distinct conditions that slow or stall replication fork elongation (replication stress2,18–20), suggesting that the recruitment of pre-replication complexes to intermittent or dormant origins might serve as redundancy to ensure the prevention of incomplete genome duplication. The flexible choice of replication origins could preserve genome integrity by coordinating replication with transcription and chromatin assembly on the shared chromatin template[14,17]

Methods

Results

Conclusion