Developmental and cancer-associated plasticity of DNA replication preferentially targets GC-poor, lowly expressed and late-replicating regions

Abstract

The spatiotemporal program of metazoan DNA replication is regulated during development and altered in cancers. We have generated novel OK-seq, Repli-seq and RNA-seq data to compare the DNA replication and gene expression programs of twelve cancer and non-cancer human cell types. Changes in replication fork directionality (RFD) determined by OK-seq are widespread but more frequent within GC-poor isochores and largely disconnected from transcription changes. Cancer cell RFD profiles cluster with non-cancer cells of similar developmental origin but not with different cancer types. Importantly, recurrent RFD changes are detected in specific tumour progression pathways. Using a model for establishment and early progression of chronic myeloid leukemia (CML), we identify 1027 replication initiation zones (IZs) that progressively change efficiency during long-term expression of the BCR-ABL1 oncogene, being twice more often downregulated than upregulated. Prolonged expression of BCR-ABL1 results in targeting of new IZs and accentuation of previous efficiency changes. Targeted IZs are predominantly located in GC-poor, late replicating gene deserts and frequently silenced in late CML. Prolonged expression of BCR-ABL1 results in massive deletion of GC-poor, late replicating DNA sequences enriched in origin silencing events. We conclude that BCR-ABL1 expression progressively affects replication and stability of GC-poor, late-replicating regions during CML progression.

Identification of new driver and passenger mutations within APOBEC-induced hotspot mutations in bladder cancer

“…Accordingly, we did not observe a coding strand bias within the 44 classified mutations (Additional file 1 : Table S1) and most of the APOBEC-associated hotspots preferentially occurred in lagging-strand templates during DNA replication compared to those non APOBEC-associated hotspot mutations (Fig. 2 a, across 9 cell lines [ 37 , 38 ], ‘ Methods ’). Additionally, DNA folding predictions indicated that the APOBEC-associated hotspots were preferentially located within the loop of DNA hairpin structures compared to non APOBEC-associated ones (Fig.…”

“…We performed analysis of published genome-wide replication fork directionality (RFD) data in nine human cancer/normal cell lines—HeLa, IMR90, TLSE19, K562, TF1, GM06990, BL79, IARC385 and Raji cells [ 37 , 38 ]—to identify the strand that would be favoured as the lagging-strand template. Considering APOBEC enzymes specifically deaminated ‘C’ to ‘U’, we expected the complementary strand to be the lagging-strand template if mutations were of the N G A → N [ C / A ] A type.…”

“…Considering APOBEC enzymes specifically deaminated ‘C’ to ‘U’, we expected the complementary strand to be the lagging-strand template if mutations were of the N G A → N [ C / A ] A type. Data availability and interpretation have been described elsewhere [ 4 , 37 , 38 ]. RFD profiles were determined by mapping Okazaki fragments to C (Crick) and W (Watson) DNA strands.…”

Active transcription regulates ORC/MCM distribution whereas replication timing correlates with ORC density in human cells

“…After confirming the validity of the ChIP experiments and establishing an analysis approach based on moderate averaging, we compared the relative read frequencies of each pre-RC component to active replication initiation units. Using OK-seq in Raji cells 37 , we calculated the replication fork directionality (RFD), and delineated zones of preferential replication initiation as ascending segments (AS) of the RFD profile. OK-seq does not detect single replication initiation events, but regions of preferential replication initiation (AS) 22, 24, 25 .…”

“…Raji OK-seq was recently published as part of Wu et al . and is available from the European Nucleotide Archive under accession number PRJEB25180 (see data access section) 37 . Reads > 10 nt were aligned to the human reference genome (hg19) using the BWA (version 0.7.4) software with default parameters 60 .…”