Research

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21 pages, 1763 KiB

Open AccessArticle

Proteins Marking the Sequence of Genotoxic Signaling from Irradiated Mesenchymal Stromal Cells to CD34+ Cells

by Vanessa Kohl, Oliver Drews, Victor Costina, Miriam Bierbaum, Ahmed Jawhar, Henning Roehl, Christel Weiss, Susanne Brendel, Helga Kleiner, Johanna Flach, Birgit Spiess, Wolfgang Seifarth, Daniel Nowak, Wolf-Karsten Hofmann, Alice Fabarius and Henning D. Popp

Int. J. Mol. Sci. 2021, 22(11), 5844; https://doi.org/10.3390/ijms22115844 - 29 May 2021

Cited by 2 | Viewed by 2770

Abstract

Non-targeted effects (NTE) of ionizing radiation may initiate myeloid neoplasms (MN). Here, protein mediators (I) in irradiated human mesenchymal stromal cells (MSC) as the NTE source, (II) in MSC conditioned supernatant and (III) in human bone marrow CD34+ cells undergoing genotoxic NTE were [...] Read more.

Non-targeted effects (NTE) of ionizing radiation may initiate myeloid neoplasms (MN). Here, protein mediators (I) in irradiated human mesenchymal stromal cells (MSC) as the NTE source, (II) in MSC conditioned supernatant and (III) in human bone marrow CD34+ cells undergoing genotoxic NTE were investigated. Healthy sublethal irradiated MSC showed significantly increased levels of reactive oxygen species. These cells responded by increasing intracellular abundance of proteins involved in proteasomal degradation, protein translation, cytoskeleton dynamics, nucleocytoplasmic shuttling, and those with antioxidant activity. Among the increased proteins were THY1 and GNA11/14, which are signaling proteins with hitherto unknown functions in the radiation response and NTE. In the corresponding MSC conditioned medium, the three chaperones GRP78, CALR, and PDIA3 were increased. Together with GPI, these were the only four altered proteins, which were associated with the observed genotoxic NTE. Healthy CD34+ cells cultured in MSC conditioned medium suffered from more than a six-fold increase in γH2AX focal staining, indicative for DNA double-strand breaks, as well as numerical and structural chromosomal aberrations within three days. At this stage, five proteins were altered, among them IQGAP1, HMGB1, and PA2G4, which are involved in malign development. In summary, our data provide novel insights into three sequential steps of genotoxic signaling from irradiated MSC to CD34+ cells, implicating that induced NTE might initiate the development of MN. Full article

(This article belongs to the Special Issue DNA Replication Stress and Chromosomal Instability)

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21 pages, 3367 KiB

Open AccessArticle

Low Replicative Stress Triggers Cell-Type Specific Inheritable Advanced Replication Timing

by Lilas Courtot, Elodie Bournique, Chrystelle Maric, Laure Guitton-Sert, Miguel Madrid-Mencía, Vera Pancaldi, Jean-Charles Cadoret, Jean-Sébastien Hoffmann and Valérie Bergoglio

Int. J. Mol. Sci. 2021, 22(9), 4959; https://doi.org/10.3390/ijms22094959 - 07 May 2021

Cited by 3 | Viewed by 3631

Abstract

DNA replication timing (RT), reflecting the temporal order of origin activation, is known as a robust and conserved cell-type specific process. Upon low replication stress, the slowing of replication forks induces well-documented RT delays associated to genetic instability, but it can also generate [...] Read more.

DNA replication timing (RT), reflecting the temporal order of origin activation, is known as a robust and conserved cell-type specific process. Upon low replication stress, the slowing of replication forks induces well-documented RT delays associated to genetic instability, but it can also generate RT advances that are still uncharacterized. In order to characterize these advanced initiation events, we monitored the whole genome RT from six independent human cell lines treated with low doses of aphidicolin. We report that RT advances are cell-type-specific and involve large heterochromatin domains. Importantly, we found that some major late to early RT advances can be inherited by the unstressed next-cellular generation, which is a unique process that correlates with enhanced chromatin accessibility, as well as modified replication origin landscape and gene expression in daughter cells. Collectively, this work highlights how low replication stress may impact cellular identity by RT advances events at a subset of chromosomal domains. Full article

(This article belongs to the Special Issue DNA Replication Stress and Chromosomal Instability)

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20 pages, 5528 KiB

Open AccessArticle

PARP Inhibition Increases the Reliance on ATR/CHK1 Checkpoint Signaling Leading to Synthetic Lethality—An Alternative Treatment Strategy for Epithelial Ovarian Cancer Cells Independent from HR Effectiveness

by Patrycja Gralewska, Arkadiusz Gajek, Agnieszka Marczak, Michał Mikuła, Jerzy Ostrowski, Agnieszka Śliwińska and Aneta Rogalska

Int. J. Mol. Sci. 2020, 21(24), 9715; https://doi.org/10.3390/ijms21249715 - 19 Dec 2020

Cited by 19 | Viewed by 5133

Abstract

Poly (ADP-ribose) polymerase inhibitor (PARPi, olaparib) impairs the repair of DNA single-strand breaks (SSBs), resulting in double-strand breaks (DSBs) that cannot be repaired efficiently in homologous recombination repair (HRR)-deficient cancers such as BRCA1/2-mutant cancers, leading to synthetic lethality. Despite the efficacy of olaparib [...] Read more.

Poly (ADP-ribose) polymerase inhibitor (PARPi, olaparib) impairs the repair of DNA single-strand breaks (SSBs), resulting in double-strand breaks (DSBs) that cannot be repaired efficiently in homologous recombination repair (HRR)-deficient cancers such as BRCA1/2-mutant cancers, leading to synthetic lethality. Despite the efficacy of olaparib in the treatment of BRCA1/2 deficient tumors, PARPi resistance is common. We hypothesized that the combination of olaparib with anticancer agents that disrupt HRR by targeting ataxia telangiectasia and Rad3-related protein (ATR) or checkpoint kinase 1 (CHK1) may be an effective strategy to reverse ovarian cancer resistance to olaparib. Here, we evaluated the effect of olaparib, the ATR inhibitor AZD6738, and the CHK1 inhibitor MK8776 alone and in combination on cell survival, colony formation, replication stress response (RSR) protein expression, DNA damage, and apoptotic changes in BRCA2 mutated (PEO-1) and HRR-proficient BRCA wild-type (SKOV-3 and OV-90) cells. Combined treatment caused the accumulation of DNA DSBs. PARP expression was associated with sensitivity to olaparib or inhibitors of RSR. Synergistic effects were weaker when olaparib was combined with CHK1i and occurred regardless of the BRCA2 status of tumor cells. Because PARPi increases the reliance on ATR/CHK1 for genome stability, the combination of PARPi with ATR inhibition suppressed ovarian cancer cell growth independently of the efficacy of HRR. The present results were obtained at sub-lethal doses, suggesting the potential of these inhibitors as monotherapy as well as in combination with olaparib. Full article

(This article belongs to the Special Issue DNA Replication Stress and Chromosomal Instability)

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Review

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21 pages, 19417 KiB

Open AccessReview

DNA Damage Response in the Adaptive Arm of the Immune System: Implications for Autoimmunity

by Theodora Manolakou, Panayotis Verginis and Dimitrios T. Boumpas

Int. J. Mol. Sci. 2021, 22(11), 5842; https://doi.org/10.3390/ijms22115842 - 29 May 2021

Cited by 5 | Viewed by 3891

Abstract

In complex environments, cells have developed molecular responses to confront threats against the genome and achieve the maintenance of genomic stability assuring the transfer of undamaged DNA to their progeny. DNA damage response (DDR) mechanisms may be activated upon genotoxic or environmental agents, [...] Read more.

In complex environments, cells have developed molecular responses to confront threats against the genome and achieve the maintenance of genomic stability assuring the transfer of undamaged DNA to their progeny. DNA damage response (DDR) mechanisms may be activated upon genotoxic or environmental agents, such as cytotoxic drugs or ultraviolet (UV) light, and during physiological processes requiring DNA transactions, to restore DNA alterations that may cause cellular malfunction and affect viability. In addition to the DDR, multicellular organisms have evolved specialized immune cells to respond and defend against infections. Both adaptive and innate immune cells are subjected to DDR processes, either as a prerequisite to the immune response, or as a result of random endogenous and exogenous insults. Aberrant DDR activities have been extensively studied in the immune cells of the innate arm, but not in adaptive immune cells. Here, we discuss how the aberrant DDR may lead to autoimmunity, with emphasis on the adaptive immune cells and the potential of therapeutic targeting. Full article

(This article belongs to the Special Issue DNA Replication Stress and Chromosomal Instability)

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36 pages, 2107 KiB

Open AccessReview

SUMO-Targeted Ubiquitin Ligases and Their Functions in Maintaining Genome Stability

by Ya-Chu Chang, Marissa K. Oram and Anja-Katrin Bielinsky

Int. J. Mol. Sci. 2021, 22(10), 5391; https://doi.org/10.3390/ijms22105391 - 20 May 2021

Cited by 23 | Viewed by 5806

Abstract

Small ubiquitin-like modifier (SUMO)-targeted E3 ubiquitin ligases (STUbLs) are specialized enzymes that recognize SUMOylated proteins and attach ubiquitin to them. They therefore connect the cellular SUMOylation and ubiquitination circuits. STUbLs participate in diverse molecular processes that span cell cycle regulated events, including DNA [...] Read more.

Small ubiquitin-like modifier (SUMO)-targeted E3 ubiquitin ligases (STUbLs) are specialized enzymes that recognize SUMOylated proteins and attach ubiquitin to them. They therefore connect the cellular SUMOylation and ubiquitination circuits. STUbLs participate in diverse molecular processes that span cell cycle regulated events, including DNA repair, replication, mitosis, and transcription. They operate during unperturbed conditions and in response to challenges, such as genotoxic stress. These E3 ubiquitin ligases modify their target substrates by catalyzing ubiquitin chains that form different linkages, resulting in proteolytic or non-proteolytic outcomes. Often, STUbLs function in compartmentalized environments, such as the nuclear envelope or kinetochore, and actively aid in nuclear relocalization of damaged DNA and stalled replication forks to promote DNA repair or fork restart. Furthermore, STUbLs reside in the same vicinity as SUMO proteases and deubiquitinases (DUBs), providing spatiotemporal control of their targets. In this review, we focus on the molecular mechanisms by which STUbLs help to maintain genome stability across different species. Full article

(This article belongs to the Special Issue DNA Replication Stress and Chromosomal Instability)

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12 pages, 715 KiB

Open AccessReview

Regulation of DNA Replication Licensing and Re-Replication by Cdt1

by Hui Zhang

Int. J. Mol. Sci. 2021, 22(10), 5195; https://doi.org/10.3390/ijms22105195 - 14 May 2021

Cited by 13 | Viewed by 3471

Abstract

In eukaryotic cells, DNA replication licensing is precisely regulated to ensure that the initiation of genomic DNA replication in S phase occurs once and only once for each mitotic cell division. A key regulatory mechanism by which DNA re-replication is suppressed is the [...] Read more.

In eukaryotic cells, DNA replication licensing is precisely regulated to ensure that the initiation of genomic DNA replication in S phase occurs once and only once for each mitotic cell division. A key regulatory mechanism by which DNA re-replication is suppressed is the S phase-dependent proteolysis of Cdt1, an essential replication protein for licensing DNA replication origins by loading the Mcm2-7 replication helicase for DNA duplication in S phase. Cdt1 degradation is mediated by CRL4^Cdt2 ubiquitin E3 ligase, which further requires Cdt1 binding to proliferating cell nuclear antigen (PCNA) through a PIP box domain in Cdt1 during DNA synthesis. Recent studies found that Cdt2, the specific subunit of CRL4^Cdt2 ubiquitin E3 ligase that targets Cdt1 for degradation, also contains an evolutionarily conserved PIP box-like domain that mediates the interaction with PCNA. These findings suggest that the initiation and elongation of DNA replication or DNA damage-induced repair synthesis provide a novel mechanism by which Cdt1 and CRL4^Cdt2 are both recruited onto the trimeric PCNA clamp encircling the replicating DNA strands to promote the interaction between Cdt1 and CRL4^Cdt2. The proximity of PCNA-bound Cdt1 to CRL4^Cdt2 facilitates the destruction of Cdt1 in response to DNA damage or after DNA replication initiation to prevent DNA re-replication in the cell cycle. CRL4^Cdt2 ubiquitin E3 ligase may also regulate the degradation of other PIP box-containing proteins, such as CDK inhibitor p21 and histone methylase Set8, to regulate DNA replication licensing, cell cycle progression, DNA repair, and genome stability by directly interacting with PCNA during DNA replication and repair synthesis. Full article

(This article belongs to the Special Issue DNA Replication Stress and Chromosomal Instability)

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26 pages, 1541 KiB

Open AccessReview

Replication Stress, Genomic Instability, and Replication Timing: A Complex Relationship

by Lina-Marie Briu, Chrystelle Maric and Jean-Charles Cadoret

Int. J. Mol. Sci. 2021, 22(9), 4764; https://doi.org/10.3390/ijms22094764 - 30 Apr 2021

Cited by 12 | Viewed by 3694

Abstract

The replication-timing program constitutes a key element of the organization and coordination of numerous nuclear processes in eukaryotes. This program is established at a crucial moment in the cell cycle and occurs simultaneously with the organization of the genome, thus indicating the vital [...] Read more.

The replication-timing program constitutes a key element of the organization and coordination of numerous nuclear processes in eukaryotes. This program is established at a crucial moment in the cell cycle and occurs simultaneously with the organization of the genome, thus indicating the vital significance of this process. With recent technological achievements of high-throughput approaches, a very strong link has been confirmed between replication timing, transcriptional activity, the epigenetic and mutational landscape, and the 3D organization of the genome. There is also a clear relationship between replication stress, replication timing, and genomic instability, but the extent to which they are mutually linked to each other is unclear. Recent evidence has shown that replication timing is affected in cancer cells, although the cause and consequence of this effect remain unknown. However, in-depth studies remain to be performed to characterize the molecular mechanisms of replication-timing regulation and clearly identify different cis- and trans-acting factors. The results of these studies will potentially facilitate the discovery of new therapeutic pathways, particularly for personalized medicine, or new biomarkers. This review focuses on the complex relationship between replication timing, replication stress, and genomic instability. Full article

(This article belongs to the Special Issue DNA Replication Stress and Chromosomal Instability)

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24 pages, 1695 KiB

Open AccessReview

DNA2 in Chromosome Stability and Cell Survival—Is It All about Replication Forks?

by Jessica J. R. Hudson and Ulrich Rass

Int. J. Mol. Sci. 2021, 22(8), 3984; https://doi.org/10.3390/ijms22083984 - 13 Apr 2021

Cited by 7 | Viewed by 3579

Abstract

The conserved nuclease-helicase DNA2 has been linked to mitochondrial myopathy, Seckel syndrome, and cancer. Across species, the protein is indispensable for cell proliferation. On the molecular level, DNA2 has been implicated in DNA double-strand break (DSB) repair, checkpoint activation, Okazaki fragment processing (OFP), [...] Read more.

The conserved nuclease-helicase DNA2 has been linked to mitochondrial myopathy, Seckel syndrome, and cancer. Across species, the protein is indispensable for cell proliferation. On the molecular level, DNA2 has been implicated in DNA double-strand break (DSB) repair, checkpoint activation, Okazaki fragment processing (OFP), and telomere homeostasis. More recently, a critical contribution of DNA2 to the replication stress response and recovery of stalled DNA replication forks (RFs) has emerged. Here, we review the available functional and phenotypic data and propose that the major cellular defects associated with DNA2 dysfunction, and the links that exist with human disease, can be rationalized through the fundamental importance of DNA2-dependent RF recovery to genome duplication. Being a crucial player at stalled RFs, DNA2 is a promising target for anti-cancer therapy aimed at eliminating cancer cells by replication-stress overload. Full article

(This article belongs to the Special Issue DNA Replication Stress and Chromosomal Instability)

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