Multi-omic analysis of altered transcriptome and epigenetic signatures in the UV-induced DNA damage response
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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2023, Current Opinion in Cell BiologyHistone H2Bub dynamics in the 5′ region of active genes are tightly linked to the UV-induced transcriptional response
2023, Computational and Structural Biotechnology JournalCitation Excerpt :Chromatin remodelling has been shown to directly affect the efficiency of repair by either of the two NER sub-pathways during the recovery of UV irradiation [40–43]. Increased chromatin accessibility [13,44] at the transcription start sites (TSSs) of active regulatory and gene regions has been reported to occur in parallel with the maintenance of active histone marks (such as histone H3K27acetylation) and the lack of deposition of transcriptional silencing modifications (H3K27me3) at the same loci in response to UV-induced genotoxic stress. Interestingly, chromatin accessibility gain is correlated with the enhanced PPP release of RNAPII into productive elongation, the continuous initiation of RNAPII from all actively transcribed regions and the accelerated repair of transcription-blocking lesions in response to UV irradiation [13].
DNMT3A-mediated high expression of circ_0057504 promotes benzo[a]pyrene-induced DNA damage via the NONO-SFPQ complex in human bronchial epithelial cells
2022, Environment InternationalCitation Excerpt :Alternative splicing and post-translational modifications play a key role in the processing of RNAs, and since circRNAs and mRNA are both derived from pre-mRNA, these modifications are important in abnormal expression of circRNAs. Previous reports have suggested that B[a]P exposure can modify genomic methylation levels (Corrales et al., 2014; Sadikovic et al., 2007), while others have shown that hypomethylation is mediated by the DDR (Feinberg et al., 2002; Liu et al., 2021). Here, we found that B[a]P treatment led to reduced mRNA and protein expression levels of DNMT3A, a de novo methyltransferase, as measured by RNA microarray, qRT-PCR and western blot assays.
Transcription and genome integrity
2022, DNA RepairCitation Excerpt :UV light induces pyrimidine dimers and 6–4 photoproducts which both efficiently block elongation of RNAPII both in vitro [29] and in vivo [30–32] (Fig. 1B). UV light causes transient decondensation of chromatin [33–38] which may reflect a cellular response that allows DNA repair enzymes to gain access to transcription-blocking lesions. While UV light exposure does not directly result in DNA strand breaks, induction of γH2AX is observed following UV irradiation in response to DNA repair-induced intermediates [39].
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These authors contributed equally.