Elsevier

DNA Repair

Volume 106, October 2021, 103172
DNA Repair

Multi-omic analysis of altered transcriptome and epigenetic signatures in the UV-induced DNA damage response

https://doi.org/10.1016/j.dnarep.2021.103172Get rights and content

Highlights

  • Increased chromatin accessibility in the early phase of DNA damage repair.

  • RNA m6A hypomethylation in response to UV treatment.

  • Loss of DNA 5mC modifications on the rapid response genes.

  • UV-induced dynamic transcriptome profiles are regulated by multifaceted epigenetic programs.

Abstract

The transcription-related DNA damage response was visualized on a genome-wide scale with great spatial and temporal resolution. Upon UV irradiation, a small proportion of mature RNA transcripts undergo changes, with significant activation of DNA repair factors. Notably, an increase of chromatin accessibility is observed at the immediate early recovery phase and serves as binding sites for selective stage-specific transcription factors. Whole genome analysis of DNA methylation (5mC) delineates pervasive dynamics during DNA repair process, and hypomethylation at gene bodies and 3’UTR is accompanied by induction of DNA damage response genes. Furthermore, temporal-specific m6A RNA methylation has been defined and appears to affect DNA repair by modulation of translation. These findings provide a resource for identifying players required for transcription-coupled nucleotide excision repair and reveal insights into the epigenetic regulation of the transcriptional programs in response to genotoxic stress.

Introduction

Transcription-coupled nucleotide excision repair (TC-NER) is promptly triggered by “local” elongating Pol II molecules encountering DNA adducts such as cyclobutane pyrimidine dimers (CPD) induced by ultraviolet (UV) light. This speeds up removal and repair of DNA lesions in transcribed strand [1,2]. Genome-wide measurements have revealed a dramatic shutdown of global transcription with a few exceptions of individual genes that are consequently highly upregulated by DNA damage [3,4]. For instances, CDKN1A is induced to elicit DNA damage-dependent cell cycle arrest [5]. Transcription factor ATF3 is immediately upregulated after DNA damage to orchestrate transcription shutdown [6]. Deciphering the underlying mechanisms of the multifaceted transcriptional response still represents one of the most fascinating frontiers in DNA repair research. Recent reports have revealed that persistent Pol II initiation at the transcription start site (TSS) of active regulatory regions with the continuous synthesis of start-RNAs guarantee sufficient scanning and repair of the whole transcribed genome, although transcription elongation drastically slows down shortly after genotoxic stress [7,8]. Furthermore, a recent study found that RNA is extremely rapidly m6A methylated by METTL3 in response to UV irradiation, which is crucial for recruitment of Pol κ to DNA damage sites and subsequently mediates TC-NER [9]. Together, these findings substantiate the claims that the molecular processes underlying the transcription-coordinated cellular response upon genotoxic stress might be more complex than previously believed. Moreover, all these findings merely focused on the early recovery phase (within 4 h in response to genotoxic stress). It remains to be determined how accessible chromatin reconfigures and regulates transcriptional programs in the whole UV-induced DNA damage repair (DDR) process. It is also unclear whether the dynamics of gene expression are associated with other epigenomic reconstruction events such as global DNA methylation and demethylation. Moreover, the roles of internal messenger RNA modifications like N6-methyladenosine in transcriptional response to DNA damage are poorly understood. As described below, our integrative analyses provide a comprehensive view of the spatiotemporal chromatin configuration that accompanies TC-NER.

Section snippets

UV-induced dynamic transcriptome profiling

In order to illustrate the transcriptomic atlas during the DDR process, we performed RNA-seq from UV-irradiated human fibroblast cell line MRC5, which have been historically used to study DNA damage and repair (Figure S1A) [10]. We collected two replicates at 3 timepoints upon UV irradiation (12 J/m2, with cell survival rate of > 50 %) that covered the key events: transcription shutdown (30 min after UV exposure, by the time the release of promoter-proximal Pol II is increased along with

Discussion

UV-induced DNA damage in mammalian cells triggers a multipronged DNA damage response, encompassing activation of DNA repair that removes lesions from the double helix, arrest of the cell-cycle progression to prevent the transmission of damaged DNA to daughter cells, stimulation of the ubiquitin-proteasome system and apoptosis that eliminate cells with heavily damaged genomes, and the transcription response that changes the cellular RNA profile. Among these four aspects, genome-wide

Cell culture

Human fibroblast cells MRC5 were cultured in DMEM containing 10 % FBS and 1% penicillin/streptomycin. 12 J/m2 UV-C was used to irradiate cells in this study.

RT-PCR

Total RNA was extracted by using an RNeasy Mini Kit (Qiagen), according to the manufacturer’s instructions. The integrity of RNA was tested on a denaturing agarose gel. RNA quality and quantity were also determined with a Nanodrop spectrophotometer (Thermo Fisher Scientific). For quantitative RT-PCR analysis, single-stranded cDNA was

Data access

All RNA-Seq, ATAC-Seq, RRBS-Seq and meRIP-Seq data used in this study are available under GEO: GSE161793.

Author contributions

J.L. and Y.W. conceived the project. J.L. performed all experiments. L.L., J.H. and Y.X. performed the bioinformatic analysis. Y.W. wrote the manuscript, with input from all authors.

Sources of funding

National Natural Science Foundation of China, Grant number: 81571092.

The Project of Department of Education of Guangdong Province of China, Grant number: 2020KTSCX099.

The Project of Guangzhou Municipal Science and Technology Bureau, Grant number: 202102080216.

The Talent Training Program of the Basic Medical College of Guangzhou Medical University, Grant number: JCXKJS2021B01.

The Science and Technology Planning Project of Guangzhou, Grant number: 201707010319.

Declaration of Competing Interest

The authors report no declarations of interest.

Acknowledgements

This work was supported by the National Natural Science Foundation of China [grant number 81571092], the Project of Department of Education of Guangdong Province of China [grant number 2020KTSCX099], the Project of Guangzhou Municipal Science and Technology Bureau [grant number 202102080216], and the Talent Training Program of the Basic Medical College of Guangzhou Medical University [grant number JCXKJS2021B01] to Y.W., and by a grant to L.L. from the Science and Technology Planning Project of

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