The type and yield of ionising radiation induced chromosomal aberrations depend on the efficiency of different DSB repair pathways in mammalian cells

Mutations in more than 200 retina-specific genes have been associated with inherited retinal diseases. Genome editing represents a promising emerging field in the treatment of monogenic disorders, as it aims to correct disease-causing mutations within the genome. Genome editing relies on highly specific endonucleases and the capacity of the cells to repair double-strand breaks (DSBs). As DSB pathways are cell-cycle dependent, their activity in postmitotic retinal neurons, with a focus on photoreceptors, needs to be assessed in order to develop therapeutic in vivo genome editing. Three DSB-repair pathways are found in mammalian cells: Non-homologous end joining (NHEJ); microhomology-mediated end joining (MMEJ); and homology-directed repair (HDR). While NHEJ can be used to knock out mutant alleles in dominant disorders, HDR and MMEJ are better suited for precise genome editing, or for replacing entire mutation hotspots in genomic regions. Here, we analyzed transcriptomic in vivo and in vitro data and revealed that HDR is indeed downregulated in postmitotic neurons, whereas MMEJ and NHEJ are active. Using single-cell RNA sequencing analysis, we characterized the dynamics of DSB repair pathways in the transition from dividing cells to postmitotic retinal cells. Time-course bulk RNA-seq data confirmed DSB repair gene expression in both in vivo and in vitro samples. Transcriptomic DSB repair pathway profiles are very similar in adult human, macaque, and mouse retinas, but not in ground squirrel retinas. Moreover, human-induced pluripotent stem-cell-derived neurons and retinal organoids can serve as well suited in vitro testbeds for developing genomic engineering approaches in photoreceptors. Our study provides additional support for designing precise in vivo genome-editing approaches via MMEJ, which is active in mature photoreceptors.

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The Determinant of DNA Repair Pathway Choices in Ionising Radiation-Induced DNA Double-Strand Breaks

BioMed Research International ◽

10.1155/2020/4834965 ◽

2020 ◽

Vol 2020 ◽

pp. 1-12 ◽

Cited By ~ 3

Author(s):

Lei Zhao ◽

Chengyu Bao ◽

Yuxuan Shang ◽

Xinye He ◽

Chiyuan Ma ◽

...

Keyword(s):

Dna Repair ◽

Double Strand Breaks ◽

Ionising Radiation ◽

Repair Pathway ◽

Dna Double Strand Breaks ◽

End Joining ◽

Dsb Repair ◽

Strand Breaks ◽

Dna Dsbs ◽

Repair Pathways

Ionising radiation- (IR-) induced DNA double-strand breaks (DSBs) are considered to be the deleterious DNA lesions that pose a serious threat to genomic stability. The major DNA repair pathways, including classical nonhomologous end joining, homologous recombination, single-strand annealing, and alternative end joining, play critical roles in countering and eliciting IR-induced DSBs to ensure genome integrity. If the IR-induced DNA DSBs are not repaired correctly, the residual or incorrectly repaired DSBs can result in genomic instability that is associated with certain human diseases. Although many efforts have been made in investigating the major mechanisms of IR-induced DNA DSB repair, it is still unclear what determines the choices of IR-induced DNA DSB repair pathways. In this review, we discuss how the mechanisms of IR-induced DSB repair pathway choices can operate in irradiated cells. We first briefly describe the main mechanisms of the major DNA DSB repair pathways and the related key repair proteins. Based on our understanding of the characteristics of IR-induced DNA DSBs and the regulatory mechanisms of DSB repair pathways in irradiated cells and recent advances in this field, We then highlight the main factors and associated challenges to determine the IR-induced DSB repair pathway choices. We conclude that the type and distribution of IR-induced DSBs, chromatin state, DNA-end structure, and DNA-end resection are the main determinants of the choice of the IR-induced DNA DSB repair pathway.

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How cancer cells hijack DNA double-strand break repair pathways to gain genomic instability

Biochemical Journal ◽

10.1042/bj20150582 ◽

2015 ◽

Vol 471 (1) ◽

pp. 1-11 ◽

Cited By ~ 48

Author(s):

Penny A. Jeggo ◽

Markus Löbrich

Keyword(s):

Cancer Cells ◽

Genomic Instability ◽

Mammalian Cells ◽

Tumour Progression ◽

Genetic Material ◽

Strand Break ◽

Cell Cycle Checkpoint ◽

Dsb Repair ◽

Normal Cells ◽

Repair Pathways

DNA DSBs (double-strand breaks) are a significant threat to the viability of a normal cell, since they can result in loss of genetic material if mitosis or replication is attempted in their presence. Consequently, evolutionary pressure has resulted in multiple pathways and responses to enable DSBs to be repaired efficiently and faithfully. Cancer cells, which are under pressure to gain genomic instability, have a striking ability to avoid the elegant mechanisms by which normal cells maintain genomic stability. Current models suggest that, in normal cells, DSB repair occurs in a hierarchical manner that promotes rapid and efficient rejoining first, with the utilization of additional steps or pathways of diminished accuracy if rejoining is unsuccessful or delayed. In the present review, we evaluate the fidelity of DSB repair pathways and discuss how cancer cells promote the utilization of less accurate processes. Homologous recombination serves to promote accuracy and stability during replication, providing a battlefield for cancer to gain instability. Non-homologous end-joining, a major DSB repair pathway in mammalian cells, usually operates with high fidelity and only switches to less faithful modes if timely repair fails. The transition step is finely tuned and provides another point of attack during tumour progression. In addition to DSB repair, a DSB signalling response activates processes such as cell cycle checkpoint arrest, which enhance the possibility of accurate DSB repair. We consider the ways by which cancers modify and hijack these processes to gain genomic instability.

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Influence of Radiation-induced Chromosomal Aberrations on the Viability of Tumour and Normal Mammalian Cells Method of Biological Dosimetry

International Journal of Radiation Biology and Related Studies in Physics Chemistry and Medicine ◽

10.1080/09553006314550431 ◽

1963 ◽

Vol 6 (4) ◽

pp. 305-322 ◽

Cited By ~ 5

Author(s):

I.M. Shapiro ◽

Z.N. Faleyeva ◽

I.B. Smirnova

Keyword(s):

Chromosomal Aberrations ◽

Mammalian Cells ◽

Biological Dosimetry ◽

Radiation Induced

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Shelterin-Mediated Telomere Protection

Annual Review of Genetics ◽

10.1146/annurev-genet-032918-021921 ◽

2018 ◽

Vol 52 (1) ◽

pp. 223-247 ◽

Cited By ~ 158

Author(s):

Titia de Lange

Keyword(s):

Dna Damage ◽

Ataxia Telangiectasia ◽

Mammalian Cells ◽

Current Knowledge ◽

Strand Break ◽

End Joining ◽

Dsb Repair ◽

Homology Directed Repair ◽

Telomere Protection ◽

Repair Pathways

For more than a decade, it has been known that mammalian cells use shelterin to protect chromosome ends. Much progress has been made on the mechanism by which shelterin prevents telomeres from inadvertently activating DNA damage signaling and double-strand break (DSB) repair pathways. Shelterin averts activation of three DNA damage response enzymes [the ataxia-telangiectasia-mutated (ATM) and ataxia telangiectasia and Rad3-related (ATR) kinases and poly(ADP-ribose) polymerase 1 (PARP1)], blocks three DSB repair pathways [classical nonhomologous end joining (c-NHEJ), alternative (alt)-NHEJ, and homology-directed repair (HDR)], and prevents hyper-resection at telomeres. For several of these functions, mechanistic insights have emerged. In addition, much has been learned about how shelterin maintains the telomeric 3′ overhang, forms and protects the t-loop structure, and promotes replication through telomeres. These studies revealed that shelterin is compartmentalized, with individual subunits dedicated to distinct aspects of the end-protection problem. This review focuses on the current knowledge of shelterin-mediated telomere protection, highlights differences between human and mouse shelterin, and discusses some of the questions that remain.

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Regulatory and Functional Involvement of Long Non-Coding RNAs in DNA Double-Strand Break Repair Mechanisms

Cells ◽

10.3390/cells10061506 ◽

2021 ◽

Vol 10 (6) ◽

pp. 1506

Author(s):

Angelos Papaspyropoulos ◽

Nefeli Lagopati ◽

Ioanna Mourkioti ◽

Andriani Angelopoulou ◽

Spyridon Kyriazis ◽

...

Keyword(s):

Therapeutic Potential ◽

Strand Break ◽

Dna Double Strand Breaks ◽

End Joining ◽

Dsb Repair ◽

Repair Mechanisms ◽

Non Coding Rnas ◽

Repair Pathways ◽

Non Homologous End Joining ◽

Living Organisms

Protection of genome integrity is vital for all living organisms, particularly when DNA double-strand breaks (DSBs) occur. Eukaryotes have developed two main pathways, namely Non-Homologous End Joining (NHEJ) and Homologous Recombination (HR), to repair DSBs. While most of the current research is focused on the role of key protein players in the functional regulation of DSB repair pathways, accumulating evidence has uncovered a novel class of regulating factors termed non-coding RNAs. Non-coding RNAs have been found to hold a pivotal role in the activation of DSB repair mechanisms, thereby safeguarding genomic stability. In particular, long non-coding RNAs (lncRNAs) have begun to emerge as new players with vast therapeutic potential. This review summarizes important advances in the field of lncRNAs, including characterization of recently identified lncRNAs, and their implication in DSB repair pathways in the context of tumorigenesis.

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Use of human lymphocyte G0 PCCs to detect intra- and inter-chromosomal aberrations for early radiation biodosimetry and retrospective assessment of radiation-induced effects

PLoS ONE ◽

10.1371/journal.pone.0216081 ◽

2019 ◽

Vol 14 (5) ◽

pp. e0216081 ◽

Cited By ~ 6

Author(s):

Terri L. Ryan ◽

Antonio G. Pantelias ◽

Georgia I. Terzoudi ◽

Gabriel E. Pantelias ◽

Adayabalam S. Balajee

Keyword(s):

Chromosomal Aberrations ◽

Human Lymphocyte ◽

Retrospective Assessment ◽

Radiation Induced ◽

Early Radiation ◽

Radiation Biodosimetry

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The Saccharomyces cerevisiae RAD6 Group Is Composed of an Error-Prone and Two Error-Free Postreplication Repair Pathways

Genetics ◽

10.1093/genetics/155.4.1633 ◽

2000 ◽

Vol 155 (4) ◽

pp. 1633-1641 ◽

Cited By ~ 11

Author(s):

Wei Xiao ◽

Barbara L Chow ◽

Stacey Broomfield ◽

Michelle Hanna

Keyword(s):

Mammalian Cells ◽

Molecular Mechanisms ◽

Signal Transducer ◽

Repair Pathway ◽

Polyubiquitin Chain ◽

Postreplication Repair ◽

Translesion Dna Synthesis ◽

Radiation Repair ◽

Repair Pathways ◽

High Degree

Abstract The RAD6 postreplication repair and mutagenesis pathway is the only major radiation repair pathway yet to be extensively characterized. It has been previously speculated that the RAD6 pathway consists of two parallel subpathways, one error free and another error prone (mutagenic). Here we show that the RAD6 group genes can be exclusively divided into three rather than two independent subpathways represented by the RAD5, POL30, and REV3 genes; the REV3 pathway is largely mutagenic, whereas the RAD5 and the POL30 pathways are deemed error free. Mutants carrying characteristic mutations in each of the three subpathways are phenotypically indistinguishable from a single mutant such as rad18, which is defective in the entire RAD6 postreplication repair/tolerance pathway. Furthermore, the rad18 mutation is epistatic to all single or combined mutations in any of the above three subpathways. Our data also suggest that MMS2 and UBC13 play a key role in coordinating the response of the error-free subpathways; Mms2 and Ubc13 form a complex required for a novel polyubiquitin chain assembly, which probably serves as a signal transducer to promote both RAD5 and POL30 error-free postreplication repair pathways. The model established by this study will facilitate further research into the molecular mechanisms of postreplication repair and translesion DNA synthesis. In view of the high degree of sequence conservation of the RAD6 pathway genes among all eukaryotes, the model presented in this study may also apply to mammalian cells and predicts links to human diseases.

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