Editor’s Choice Articles

Editor’s Choice articles are based on recommendations by the scientific editors of MDPI journals from around the world. Editors select a small number of articles recently published in the journal that they believe will be particularly interesting to readers, or important in the respective research area. The aim is to provide a snapshot of some of the most exciting work published in the various research areas of the journal.

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15 pages, 1159 KiB  
Review
Evolution of Acquired Drug Resistance in BRAF-Mutant Melanoma
by Josué Ballesteros-Álvarez and Ana M. Blázquez-Medela
DNA 2024, 4(4), 355-369; https://doi.org/10.3390/dna4040025 - 12 Oct 2024
Viewed by 563
Abstract
Melanoma is a highly aggressive type of skin cancer. Metastatic melanoma tumors have historically featured a particularly poor prognosis and have often been considered incurable. Recent advances in targeted therapeutic interventions have radically changed the landscape in metastatic melanoma management, significantly increasing the [...] Read more.
Melanoma is a highly aggressive type of skin cancer. Metastatic melanoma tumors have historically featured a particularly poor prognosis and have often been considered incurable. Recent advances in targeted therapeutic interventions have radically changed the landscape in metastatic melanoma management, significantly increasing the overall survival of patients. Hyperactive BRAF is the most common mutational event found in metastatic melanoma and its inhibition has proven to be a successful approach in a number of patients. Unfortunately, initial tumor retreat is followed by relapse in most cases, highlighting the elusiveness of finding a widely effective treatment. Melanoma tumors often carry a particularly high number of mutations in what is known as a high level of inter- and intra-patient tumor heterogeneity, driving resistance to treatment. The various mutations that are present in these tumors, in addition to impacting the root cause of the malignancy and the potential for therapeutic interventions, have also been known to arise during tumor clonal evolution leading to the establishment of drug resistance, a major issue in melanoma management. Full article
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10 pages, 4398 KiB  
Article
Deregulation and Shattering of Chromosomal Segments Containing Multiple Oncogenic Targets in the Pathogenesis of Diffuse Large B Cell Lymphoma, Not Otherwise Specified (DLBCL, NOS)
by Ashwini K. Yenamandra, Rebecca B. Smith, Adam C. Seegmiller, Brianna N. Smith, Debra L. Friedman and Christine M. Smith
DNA 2024, 4(3), 318-327; https://doi.org/10.3390/dna4030021 - 18 Sep 2024
Viewed by 947
Abstract
Diffuse large B cell lymphoma, not otherwise specified (DLBCL, NOS) is the most common type of non-Hodgkin lymphoma (NHL). Significant efforts have been focused on utilizing advanced genomic technologies to further subclassify DLBCL, NOS into clinically relevant subtypes. These efforts have led to [...] Read more.
Diffuse large B cell lymphoma, not otherwise specified (DLBCL, NOS) is the most common type of non-Hodgkin lymphoma (NHL). Significant efforts have been focused on utilizing advanced genomic technologies to further subclassify DLBCL, NOS into clinically relevant subtypes. These efforts have led to the implementation of novel algorithms to support optimal risk-oriented therapy and improvement in the overall survival of DLBCL patients. The pathogenesis of DLBCL at the molecular level indicates copy number variation (CNV) as one of the major forms of genetic alterations in the somatic mutational landscape. Random deregulation that results in complex breaks of chromosomes and restructuring of shattered chromosomal segments is called chromothripsis. Gene expression changes influenced by chromothripsis have been reported in cancer and congenital diseases. This chaotic phenomenon results in complex CNV, gene fusions, and amplification and loss of tumor suppressor genes. We present herein a summary of the most clinically relevant genomic aberrations, with particular focus on copy number aberrations in a case that highlights DLBCL, NOS arising from relapsed Hodgkin lymphoma. The focus of our study was to understand the relationship between the clinical, morphological, and genomic abnormalities in DLBCL, NOS through multiple techniques for therapeutic considerations. Full article
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18 pages, 1984 KiB  
Review
8-OxodG: A Potential Biomarker for Chronic Oxidative Stress Induced by High-LET Radiation
by Kamendra Kumar, Albert J. Fornace, Jr. and Shubhankar Suman
DNA 2024, 4(3), 221-238; https://doi.org/10.3390/dna4030015 - 1 Aug 2024
Viewed by 1343
Abstract
Oxidative stress-mediated biomolecular damage is a characteristic feature of ionizing radiation (IR) injury, leading to genomic instability and chronic health implications. Specifically, a dose- and linear energy transfer (LET)-dependent persistent increase in oxidative DNA damage has been reported in many tissues and biofluids [...] Read more.
Oxidative stress-mediated biomolecular damage is a characteristic feature of ionizing radiation (IR) injury, leading to genomic instability and chronic health implications. Specifically, a dose- and linear energy transfer (LET)-dependent persistent increase in oxidative DNA damage has been reported in many tissues and biofluids months after IR exposure. Contrary to low-LET photon radiation, high-LET IR exposure is known to cause significantly higher accumulations of DNA damage, even at sublethal doses, compared to low-LET IR. High-LET IR is prevalent in the deep space environment (i.e., beyond Earth’s magnetosphere), and its exposure could potentially impair astronauts’ health. Therefore, the development of biomarkers to assess and monitor the levels of oxidative DNA damage can aid in the early detection of health risks and would also allow timely intervention. Among the recognized biomarkers of oxidative DNA damage, 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-OxodG) has emerged as a promising candidate, indicative of chronic oxidative stress. It has been reported to exhibit differing levels following equivalent doses of low- and high-LET IR. This review discusses 8-OxodG as a potential biomarker of high-LET radiation-induced chronic stress, with special emphasis on its potential sources, formation, repair mechanisms, and detection methods. Furthermore, this review addresses the pathobiological implications of high-LET IR exposure and its association with 8-OxodG. Understanding the association between high-LET IR exposure-induced chronic oxidative stress, systemic levels of 8-OxodG, and their potential health risks can provide a framework for developing a comprehensive health monitoring biomarker system to safeguard the well-being of astronauts during space missions and optimize long-term health outcomes. Full article
(This article belongs to the Special Issue Physics and Chemistry of Radiation Damage to DNA and Its Consequences)
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11 pages, 1028 KiB  
Article
Efficient Elimination of mtDNA from Mammalian Cells with 2′,3′-Dideoxycytidine
by Natalya Kozhukhar and Mikhail F. Alexeyev
DNA 2024, 4(3), 201-211; https://doi.org/10.3390/dna4030013 - 4 Jul 2024
Viewed by 1276
Abstract
Mammalian cell lines devoid of mitochondrial DNA (mtDNA) are indispensable in studies aimed at elucidating the contribution of mtDNA to various cellular processes or interactions between nuclear and mitochondrial genomes. However, the repertoire of tools for generating such cells (also known as rho-0 [...] Read more.
Mammalian cell lines devoid of mitochondrial DNA (mtDNA) are indispensable in studies aimed at elucidating the contribution of mtDNA to various cellular processes or interactions between nuclear and mitochondrial genomes. However, the repertoire of tools for generating such cells (also known as rho-0 or ρ0 cells) remains limited, and approaches remain time- and labor-intensive, ultimately limiting their availability. Ethidium bromide (EtBr), which is most commonly used to induce mtDNA loss in mammalian cells, is cytostatic and mutagenic as it affects both nuclear and mitochondrial genomes. Therefore, there is growing interest in new tools for generating ρ0 cell lines. Here, we examined the utility of 2′,3′-dideoxycytidine (ddC, zalcitabine) alone or in combination with EtBr for generating ρ0 cell lines of mouse and human origin as well as inducing the ρ0 state in mouse/human somatic cell hybrids. We report that ddC is superior to EtBr in both immortalized mouse fibroblasts and human 143B cells. Also, unlike EtBr, ddC exhibits no cytostatic effects at the highest concentration tested (200 μM), making it more suitable for general use. We conclude that ddC is a promising new tool for generating mammalian ρ0 cell lines. Full article
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9 pages, 459 KiB  
Article
Child Telomere Length at 11–12 Years of Age Is Not Associated with Pregnancy Complications
by Tina Bianco-Miotto, Sadia Hossain, Nahal Habibi, Dandara G. Haag and Jessica A. Grieger
DNA 2024, 4(2), 180-188; https://doi.org/10.3390/dna4020011 - 11 Jun 2024
Cited by 1 | Viewed by 809
Abstract
Children born from pregnancy complications are at higher risk of chronic diseases in adulthood. Identifying which children born from a complicated pregnancy are likely to suffer from later chronic disease is important in order to intervene to prevent or delay the onset of [...] Read more.
Children born from pregnancy complications are at higher risk of chronic diseases in adulthood. Identifying which children born from a complicated pregnancy are likely to suffer from later chronic disease is important in order to intervene to prevent or delay the onset of disease. This study examined the associations between the major pregnancy complications (gestational diabetes, high blood pressure, small- and large for gestational age, and preterm birth) and child telomere length, a biomarker of chronic disease risk. This was a population-based longitudinal analysis using data from the Longitudinal Study of Australian Children. The primary outcome is telomere length, measured in 11–12-year-old children. Multivariable linear regression was used to estimate the association between pregnancy complications and child telomere length, adjusting for a range of a priori confounders. Data from 841 families were used. One in four pregnancies (27.1%) featured a pregnancy complication. In the adjusted analysis, there was no association between pregnancy complications and child telomere length (high blood pressure: mean difference (95% CI): 0.00 (−0.12, 0.12); gestational diabetes (0.05 (−0.10, 0.19)); small for gestational age (0.07 (−0.04, 0.19)); large for gestational age (−0.06 (−0.15, 0.03)); and preterm birth (−0.10 (−0.21, 0.01)). Our results do not support the notion that telomere length is shorter in children born to mothers after a pregnancy complication. Methodological considerations should be rigorous to improve the reproducibility of findings. Full article
(This article belongs to the Special Issue Epigenetics and Environmental Exposures)
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10 pages, 1911 KiB  
Article
Role of Supercoiling and Topoisomerases in DNA Knotting
by Jorge Cebrián, María-Luisa Martínez-Robles, Victor Martínez, Pablo Hernández, Dora B. Krimer, Jorge B. Schvartzman and María-José Fernández-Nestosa
DNA 2024, 4(2), 170-179; https://doi.org/10.3390/dna4020010 - 27 May 2024
Viewed by 1644
Abstract
DNA knots are deleterious for living cells if not removed. Several theoretical and simulation approaches address the question of how topoisomerases select the intermolecular passages that preferentially lead to unknotting rather than to the knotting of randomly fluctuating DNA molecules, but the formation [...] Read more.
DNA knots are deleterious for living cells if not removed. Several theoretical and simulation approaches address the question of how topoisomerases select the intermolecular passages that preferentially lead to unknotting rather than to the knotting of randomly fluctuating DNA molecules, but the formation of knots in vivo remains poorly understood. DNA knots form in vivo in non-replicating and replicating molecules, and supercoiling as well as intertwining are thought to play a crucial role in both the formation and resolution of DNA knots by topoisomerase IV. To confirm this idea, we used two-dimensional agarose gel electrophoresis run with different concentrations of chloroquine to demonstrate that non-replicating pBR322 plasmids grown in a topoisomerase I-defective E. coli strain (RS2λ) were more negatively supercoiled than in a wild-type strain (W3110) and, concurrently, showed significantly fewer knots. In this way, using wild-type and E. coli mutant strains, we confirmed that one of the biological functions of DNA supercoiling is to reduce the formation of DNA knots. Full article
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17 pages, 1963 KiB  
Review
Mutagenesis and Repair of γ-Radiation- and Radical-Induced Tandem DNA Lesions
by Ashis K. Basu, Laureen C. Colis and Jan Henric T. Bacurio
DNA 2024, 4(2), 154-169; https://doi.org/10.3390/dna4020009 - 6 May 2024
Cited by 1 | Viewed by 1402
Abstract
Ionizing radiation induces many different types of DNA lesions. But one of its characteristics is to produce complex DNA damage, of which tandem DNA damage has received much attention, owing to its promise of distinctive biological properties. Oxidative stresses in response to inflammation [...] Read more.
Ionizing radiation induces many different types of DNA lesions. But one of its characteristics is to produce complex DNA damage, of which tandem DNA damage has received much attention, owing to its promise of distinctive biological properties. Oxidative stresses in response to inflammation in tissues and metal-catalyzed reactions that result in generation of radicals also form these DNA lesions. In this minireview, we have summarized the formation of the tandem lesions as well as the replication and repair studies carried out on them after site-specific synthesis. Many of these lesions are resistant to the traditional base excision repair, so that they can only be repaired by the nucleotide excision repair pathway. They also block DNA replication and, when lesion bypass occurs, it may be significantly error-prone. Some of these tandem DNA lesions may contribute to ageing, neurological diseases, and cancer. Full article
(This article belongs to the Special Issue Physics and Chemistry of Radiation Damage to DNA and Its Consequences)
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20 pages, 1764 KiB  
Review
How Chromatin Motor Complexes Influence the Nuclear Architecture: A Review of Chromatin Organization, Cohesins, and Condensins with a Focus on C. elegans
by Bahaar Chawla and Györgyi Csankovszki
DNA 2024, 4(1), 84-103; https://doi.org/10.3390/dna4010005 - 11 Mar 2024
Viewed by 1849
Abstract
Chromatin is the complex of DNA and associated proteins found in the nuclei of living organisms. How it is organized is a major research field as it has implications for replication, repair, and gene expression. This review summarizes the current state of the [...] Read more.
Chromatin is the complex of DNA and associated proteins found in the nuclei of living organisms. How it is organized is a major research field as it has implications for replication, repair, and gene expression. This review summarizes the current state of the chromatin organization field, with a special focus on chromatin motor complexes cohesin and condensin. Containing the highly conserved SMC proteins, these complexes are responsible for organizing chromatin during cell division. Additionally, research has demonstrated that condensin and cohesin also have important functions during interphase to shape the organization of chromatin and regulate expression of genes. Using the model organism C. elegans, the authors review the current knowledge of how these complexes perform such diverse roles and what open questions still exist in the field. Full article
(This article belongs to the Special Issue DNA Organization in Model Organisms)
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20 pages, 1247 KiB  
Review
Chromatin Organization during C. elegans Early Development
by Eshna Jash and Györgyi Csankovszki
DNA 2024, 4(1), 64-83; https://doi.org/10.3390/dna4010004 - 22 Feb 2024
Cited by 1 | Viewed by 1647
Abstract
Embryogenesis is characterized by dynamic chromatin remodeling and broad changes in chromosome architecture. These changes in chromatin organization are accompanied by transcriptional changes, which are crucial for the proper development of the embryo. Several independent mechanisms regulate this process of chromatin reorganization, including [...] Read more.
Embryogenesis is characterized by dynamic chromatin remodeling and broad changes in chromosome architecture. These changes in chromatin organization are accompanied by transcriptional changes, which are crucial for the proper development of the embryo. Several independent mechanisms regulate this process of chromatin reorganization, including the segregation of chromatin into heterochromatin and euchromatin, deposition of active and repressive histone modifications, and the formation of 3D chromatin domains such as TADs and LADs. These changes in chromatin structure are directly linked to developmental milestones such as the loss of developmental plasticity and acquisition of terminally differentiated cell identities. In this review, we summarize these processes that underlie this chromatin reorganization and their impact on embryogenesis in the nematode C. elegans. Full article
(This article belongs to the Special Issue DNA Organization in Model Organisms)
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12 pages, 910 KiB  
Article
Comparison of Reduced PCR Volume PowerPlex Fusion 6C Kit Validations on Manual and Automated Systems
by Eszter É. Lőrincz, Norbert Mátrai, Katalin A. Rádóczy, Tamás Cseppentő, Nóra M. Magonyi and Attila Heinrich
DNA 2024, 4(1), 52-63; https://doi.org/10.3390/dna4010003 - 4 Feb 2024
Cited by 3 | Viewed by 1965
Abstract
The PowerPlex Fusion 6C PCR™ amplification kit provides a strong discriminatory power for human identification. We have validated the kit with a reduced volume (12.5 µL) and as part of the validation we compared the efficiency of the polymerase chain reaction (PCR) prepared [...] Read more.
The PowerPlex Fusion 6C PCR™ amplification kit provides a strong discriminatory power for human identification. We have validated the kit with a reduced volume (12.5 µL) and as part of the validation we compared the efficiency of the polymerase chain reaction (PCR) prepared manually and on Hamilton Microlab® Autolys STAR Biorobot. Three years of casework data has been also included in the validation. Optimisation was carried out on different types of samples (blood, saliva, semen) and DNA was extracted robotically. Tests were conducted at two different cycle numbers (30;32), followed by analysis on both the Applied BiosystemsTM 3500 and 3500 xL Genetic Analyzer instruments (Applied Biosystems®, Foster City, CA, USA). When the PCR was prepared manually, no allele dropout was observed over 0.15 ng input DNA. Whereas when the PCR was prepared robotically, dropout already appeared at the level of 0.15 ng input DNA. In cases when increased cycle number was utilised, an increasing number of dropouts started to arise from 0.075 ng total input DNA. Despite the fact that robotically prepared PCR produced more missing alleles than the manually prepared PCR, using the optimal 0.5 ng input DNA, both methods proved to be reliable. Based on the results, our half-volume protocol is robust, and after three years of application it has proven to be effective with respect to a large number of casework samples. Full article
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18 pages, 3622 KiB  
Article
The Effects of Particle LET and Fluence on the Complexity and Frequency of Clustered DNA Damage
by Mohammad Rezaee and Amitava Adhikary
DNA 2024, 4(1), 34-51; https://doi.org/10.3390/dna4010002 - 5 Jan 2024
Cited by 3 | Viewed by 1717
Abstract
Motivation: Clustered DNA-lesions are predominantly induced by ionizing radiation, particularly by high-LET particles, and considered as lethal damage. Quantification of this specific type of damage as a function of radiation parameters such as LET, dose rate, dose, and particle type can be [...] Read more.
Motivation: Clustered DNA-lesions are predominantly induced by ionizing radiation, particularly by high-LET particles, and considered as lethal damage. Quantification of this specific type of damage as a function of radiation parameters such as LET, dose rate, dose, and particle type can be informative for the prediction of biological outcome in radiobiological studies. This study investigated the induction and complexity of clustered DNA damage for three different types of particles at an LET range of 0.5–250 keV/µm. Methods: Nanometric volumes (36.0 nm3) of 15 base-pair DNA with its hydration shell was modeled. Electron, proton, and alpha particles at various energies were simulated to irradiate the nanometric volumes. The number of ionization events, low-energy electron spectra, and chemical yields for the formation of °OH, H°, eaq, and H2O2 were calculated for each particle as a function of LET. Single- and double-strand breaks (SSB and DSB), base release, and clustered DNA-lesions were computed from the Monte-Carlo based quantification of the reactive species and measured yields of the species responsible for the DNA lesion formation. Results: The total amount of DNA damage depends on particle type and LET. The number of ionization events underestimates the quantity of DNA damage at LETs higher than 10 keV/µm. Minimum LETs of 9.4 and 11.5 keV/µm are required to induce clustered damage by a single track of proton and alpha particles, respectively. For a given radiation dose, an increase in LET reduces the number of particle tracks, leading to more complex clustered DNA damage, but a smaller number of separated clustered damage sites. Conclusions: The dependency of the number and the complexity of clustered DNA damage on LET and fluence suggests that the quantification of this damage can be a useful method for the estimation of the biological effectiveness of radiation. These results also suggest that medium-LET particles are more appropriate for the treatment of bulk targets, whereas high-LET particles can be more effective for small targets. Full article
(This article belongs to the Special Issue Physics and Chemistry of Radiation Damage to DNA and Its Consequences)
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33 pages, 4701 KiB  
Article
Evaluating Metabarcoding Markers for Identifying Zooplankton and Ichthyoplankton Communities to Species in the Salish Sea: Morphological Comparisons and Rare, Threatened or Invasive Species
by Carol A. Stepien, Haila K. Schultz, Sean M. McAllister, Emily L. Norton and Julie E. Keister
DNA 2024, 4(1), 1-33; https://doi.org/10.3390/dna4010001 - 22 Dec 2023
Cited by 2 | Viewed by 2039
Abstract
Zooplankton and ichthyoplankton community assessments depend on species diagnostics, yet morphological identifications are time-consuming, require taxonomic expertise, and are hampered by a lack of diagnostic characters, particularly for larval stages. Metabarcoding can identify multiple species in communities from short DNA sequences in comparison [...] Read more.
Zooplankton and ichthyoplankton community assessments depend on species diagnostics, yet morphological identifications are time-consuming, require taxonomic expertise, and are hampered by a lack of diagnostic characters, particularly for larval stages. Metabarcoding can identify multiple species in communities from short DNA sequences in comparison to reference databases. To evaluate species resolution across phylogenetic groups and food webs of zooplankton and ichthyoplankton, we compare five metabarcode mitochondrial (mt)DNA markers from gene regions of (a) cytochrome c oxidase subunit I, (b) cytochrome b, (c) 16S ribosomal RNA, and (d) 12S ribosomal RNA for DNA extracted from net tows in the Northeastern Pacific Ocean’s Salish Sea across seven sites and two seasons. Species resolved by metabarcoding are compared to invertebrate morphological identifications and biomass estimates. Results indicate that species resolution for different zooplankton and ichthyoplankton taxa can markedly vary among gene regions and markers in comparison to morphological identifications. Thus, researchers seeking “universal” metabarcoding should take caution that several markers and gene regions likely will be needed; all will miss some taxa and yield incomplete overlap. Species resolution requires careful attention to taxon marker selection and coverage in reference sequence repositories. In summary, combined multi-marker metabarcoding and morphological approaches improve broadscale zooplankton diagnostics. Full article
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20 pages, 2024 KiB  
Article
Genetic Insights into Teratozoospermia: A Comprehensive Computational Study of UTR Variants in AURKC, SPATA16, and SUN5
by Maria-Anna Kyrgiafini and Zissis Mamuris
DNA 2023, 3(4), 148-167; https://doi.org/10.3390/dna3040013 - 26 Oct 2023
Viewed by 1768
Abstract
Teratozoospermia, a complex male fertility disorder affecting sperm morphology, has been linked to AURKC, SPATA16, and SUN5 gene defects. However, the sheer volume of SNPs in these genes necessitates prioritization for comprehensive analysis. This study focuses on the often-overlooked untranslated region [...] Read more.
Teratozoospermia, a complex male fertility disorder affecting sperm morphology, has been linked to AURKC, SPATA16, and SUN5 gene defects. However, the sheer volume of SNPs in these genes necessitates prioritization for comprehensive analysis. This study focuses on the often-overlooked untranslated region (UTR) variants in these genes, aiming to assess their association with teratozoospermia and prioritize them. We employed a multi-step filtering process, including functional significance assessment (RegulomeDB, 3DSNP v2.0, SNPinfo (FuncPred)), evaluation of gene expression impacts in testis tissue using GTEx, and assessment of miRNA binding site effects (PolymiRTS Database 3.0, miRNASNP v3). Additionally, we used SNPnexus to evaluate their conservation and association with diseases. In AURKC, we identified six UTR SNPs (rs11084490, rs58264281, rs35582299, rs533889458, rs2361127, rs55710619), two of which influenced gene expression in testis, while others affected the binding sites of 29 miRNAs or were located in transcription-factor binding sites. Three of these SNPs were also found to be associated with spermatogenic failure according to previous studies indicating a potential regulatory role in teratozoospermia, too. For SPATA16, two 3′ UTR variants, rs146640459 and rs148085657, were prioritized, with the latter impacting miRNA binding sites. In SUN5, three 3′ UTR variants (rs1485087675, rs762026146, rs1478197315) affected miRNA binding sites. It should be noted that none of the above variants was identified in a conserved region. Our findings shed light on the potential regulatory roles of these SNPs in teratozoospermia and lay the foundation for future research directions in this area. Full article
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15 pages, 2600 KiB  
Article
Disease-Associated Mutation A554V Disrupts Normal Autoinhibition of DNMT1
by Rebecca L. Switzer, Zach J. Hartman, Geoffrey R. Hewett and Clara F. Carroll
DNA 2023, 3(3), 119-133; https://doi.org/10.3390/dna3030010 - 13 Jul 2023
Viewed by 1943
Abstract
DNA methyltransferase 1 (DNMT1) is the enzyme primarily responsible for propagation of the methylation pattern in cells. Mutations in DNMT1 have been linked to the development of adult-onset neurodegenerative disorders; these disease-associated mutations occur in the regulatory replication foci-targeting sequence (RFTS) domain of [...] Read more.
DNA methyltransferase 1 (DNMT1) is the enzyme primarily responsible for propagation of the methylation pattern in cells. Mutations in DNMT1 have been linked to the development of adult-onset neurodegenerative disorders; these disease-associated mutations occur in the regulatory replication foci-targeting sequence (RFTS) domain of the protein. The RFTS domain is an endogenous inhibitor of DNMT1 activity that binds to the active site and prevents DNA binding. Here, we examine the impact of the disease-associated mutation A554V on normal RFTS-mediated inhibition of DNMT1. Wild-type and mutant proteins were expressed and purified to homogeneity for biochemical characterization. The mutation increased DNA binding affinity ~8-fold. In addition, the mutant enzyme exhibited increased DNA methylation activity. Circular dichroism (CD) spectroscopy revealed that the mutation does not significantly impact the secondary structure or relative thermal stability of the isolated RFTS domain. However, the mutation resulted in changes in the CD spectrum in the context of the larger protein; a decrease in relative thermal stability was also observed. Collectively, this evidence suggests that A554V disrupts normal RFTS-mediated autoinhibition of DNMT1, resulting in a hyperactive mutant enzyme. While the disease-associated mutation does not significantly impact the isolated RFTS domain, the mutation results in a weakening of the interdomain stabilizing interactions generating a more open, active conformation of DNMT1. Hyperactive mutant DNMT1 could be responsible for the increased DNA methylation observed in affected individuals. Full article
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16 pages, 2480 KiB  
Article
Exploration of the DNA Photocleavage Activity of O-Halo-phenyl Carbamoyl Amidoximes: Studies of the UVA-Induced Effects on a Major Crop Pest, the Whitefly Bemisia tabaci
by Anastasios Panagopoulos, Konstantina Alipranti, Kyriaki Mylona, Polinikis Paisidis, Stergios Rizos, Alexandros E. Koumbis, Emmanouil Roditakis and Konstantina C. Fylaktakidou
DNA 2023, 3(2), 85-100; https://doi.org/10.3390/dna3020006 - 4 Apr 2023
Cited by 1 | Viewed by 1950
Abstract
The DNA photocleavage effect of halogenated O-carbamoyl derivatives of 4-MeO-benzamidoxime under UVB and UVA irradiation was studied in order to identify the nature, position, and number of halogens on the carbamoyl moiety that ensure photoactivity. F, Cl, and Br-phenyl carbamate esters (PCME) [...] Read more.
The DNA photocleavage effect of halogenated O-carbamoyl derivatives of 4-MeO-benzamidoxime under UVB and UVA irradiation was studied in order to identify the nature, position, and number of halogens on the carbamoyl moiety that ensure photoactivity. F, Cl, and Br-phenyl carbamate esters (PCME) exhibited activity with the p-Cl-phenyl derivative to show excellent photocleavage against pBR322 plasmid DNA. m-Cl-PCME has diminished activity, whereas the presence of two halogen atoms reduced DNA photocleavage. The substitution on the benzamidoxime scaffold was irrelevant to the activity. The mechanism of action indicated function in the absence of oxygen, probably via radicals derived from the N-O bond homolysis of the carbamates and in air via hydroxyl radicals and partially singlet oxygen. The UVA-vis area of absorption of the nitro-benzamidoxime p-Cl-PCMEs allowed for the investigation of their potential efficacy as photopesticides under UVA irradiation against the whitefly Bemisia tabaci, a major pest of numerous crops. The m-nitro derivative exhibited a moderate specificity against the adult population. Nymphs were not affected. The compound was inactive in the dark. This result may allow for the development of lead compounds for the control of agricultural insect pests that can cause significant economic damage in crop production. Full article
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20 pages, 719 KiB  
Review
Kinetic Studies on the 2-Oxoglutarate/Fe(II)-Dependent Nucleic Acid Modifying Enzymes from the AlkB and TET Families
by Zhiyuan Peng, Jian Ma, Christo Z. Christov, Tatyana Karabencheva-Christova, Nicolai Lehnert and Deyu Li
DNA 2023, 3(2), 65-84; https://doi.org/10.3390/dna3020005 - 30 Mar 2023
Cited by 4 | Viewed by 3004
Abstract
Nucleic acid methylations are important genetic and epigenetic biomarkers. The formation and removal of these markers is related to either methylation or demethylation. In this review, we focus on the demethylation or oxidative modification that is mediated by the 2-oxoglutarate (2-OG)/Fe(II)-dependent AlkB/TET family [...] Read more.
Nucleic acid methylations are important genetic and epigenetic biomarkers. The formation and removal of these markers is related to either methylation or demethylation. In this review, we focus on the demethylation or oxidative modification that is mediated by the 2-oxoglutarate (2-OG)/Fe(II)-dependent AlkB/TET family enzymes. In the catalytic process, most enzymes oxidize 2-OG to succinate, in the meantime oxidizing methyl to hydroxymethyl, leaving formaldehyde and generating demethylated base. The AlkB enzyme from Escherichia coli has nine human homologs (ALKBH1-8 and FTO) and the TET family includes three members, TET1 to 3. Among them, some enzymes have been carefully studied, but for certain enzymes, few studies have been carried out. This review focuses on the kinetic properties of those 2-OG/Fe(II)-dependent enzymes and their alkyl substrates. We also provide some discussions on the future directions of this field. Full article
(This article belongs to the Special Issue From Mutation and Repair to Therapeutics)
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20 pages, 1279 KiB  
Review
DNA Damage and the Gut Microbiome: From Mechanisms to Disease Outcomes
by Yun-Chung Hsiao, Chih-Wei Liu, Yifei Yang, Jiahao Feng, Haoduo Zhao and Kun Lu
DNA 2023, 3(1), 13-32; https://doi.org/10.3390/dna3010002 - 1 Feb 2023
Cited by 11 | Viewed by 5099
Abstract
Both the number of cells and the collective genome of the gut microbiota outnumber their mammalian hosts, and the metabolic and physiological interactions of the gut microbiota with the host have not yet been fully characterized. Cancer remains one of the leading causes [...] Read more.
Both the number of cells and the collective genome of the gut microbiota outnumber their mammalian hosts, and the metabolic and physiological interactions of the gut microbiota with the host have not yet been fully characterized. Cancer remains one of the leading causes of death, and more research into the critical events that can lead to cancer and the importance of the gut microbiota remains to be determined. The gut microbiota can release microbial molecules that simulate host endogenous processes, such as inflammatory responses, or can alter host metabolism of ingested substances. Both of these reactions can be beneficial or deleterious to the host, and some can be genotoxic, thus contributing to cancer progression. This review focused on the molecular evidence currently available on the mechanistic understanding of how the gut microbiota are involved in human carcinogenesis. We first reviewed the key events of carcinogenesis, especially how DNA damage proceeds to tumor formulation. Then, the current knowledge on host DNA damage attributed to the gut microbiota was summarized, followed by the genotoxic endogenous processes the gut microbiota can induce. Finally, we touched base on the association between specific gut microbiota dysbiosis and different types of cancer and concluded with the up-to-date knowledge as well as future research direction for advancing our understanding of the relationship between the gut microbiota and cancer development. Full article
(This article belongs to the Special Issue From Mutation and Repair to Therapeutics)
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12 pages, 1663 KiB  
Review
DNA G-Quadruplex-Binding Proteins: An Updated Overview
by Victoria Sanchez-Martin
DNA 2023, 3(1), 1-12; https://doi.org/10.3390/dna3010001 - 11 Jan 2023
Cited by 10 | Viewed by 5263
Abstract
DNA G-quadruplexes (G4s) are non-canonical secondary structures formed in guanine-rich sequences. Within the human genome, G4s are found in regulatory regions such as gene promoters and telomeres to control replication, transcription, and telomere lengthening. In the cellular context, there are several proteins named [...] Read more.
DNA G-quadruplexes (G4s) are non-canonical secondary structures formed in guanine-rich sequences. Within the human genome, G4s are found in regulatory regions such as gene promoters and telomeres to control replication, transcription, and telomere lengthening. In the cellular context, there are several proteins named as G4-binding proteins (G4BPs) that interact with G4s, either anchoring upon, stabilizing, and/or unwinding them. These proteins may play different key roles in the regulation of the endogenous G4 landscape and its associated functions. The present review summarizes the current literature on G4BPs in terms of their targets and functions, providing updated insights into the regulation of G4s in living organisms. Full article
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12 pages, 2094 KiB  
Article
Evaluation of Thermal Stability of DNA Oligonucleotide Structures Embedded in Hydrogels
by Daisuke Yamaguchi, Masatoshi Yoshida and Shu-ichi Nakano
DNA 2022, 2(4), 302-313; https://doi.org/10.3390/dna2040021 - 14 Dec 2022
Viewed by 3649
Abstract
Understanding the self-assembly and hybridization properties of DNA oligonucleotides in confined spaces can help to improve their applications in biotechnology and nanotechnology. This study investigates the effects of spatial confinement in the pores of hydrogels on the thermal stability of DNA oligonucleotide structures. [...] Read more.
Understanding the self-assembly and hybridization properties of DNA oligonucleotides in confined spaces can help to improve their applications in biotechnology and nanotechnology. This study investigates the effects of spatial confinement in the pores of hydrogels on the thermal stability of DNA oligonucleotide structures. The preparation of oligonucleotides embedded in agarose gels was simple, whereas the preparation of oligonucleotides embedded in polyacrylamide gels was required to remove unpolymerized monomers. In the latter case, a method for rehydrating a washed dry gel with a buffer solution containing oligonucleotides was developed. Fluorescence measurements of oligonucleotides bearing fluorescent probes revealed no significant influence of the internal environment of the gel pores on the stability of DNA duplex, hairpin, and G-quadruplex structures. Moreover, the effects of poly(ethylene glycol) on the stability of DNA structures in the gels were similar to those in solutions. It is likely that the oligonucleotides are not strongly constrained in the gels and may be preferentially located in a water-rich environment in the gel matrix. The gel preparation was also applied to the assessment of the stability of DNA structures under the conditions of a reduced number of water molecules. The studies using hydrogels provide insights into the ability of self-assembly and hybridization of oligonucleotides in confined environments and under low-water-content conditions. Full article
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23 pages, 2179 KiB  
Review
Complex Roles of NEIL1 and OGG1: Insights Gained from Murine Knockouts and Human Polymorphic Variants
by R. Stephen Lloyd
DNA 2022, 2(4), 279-301; https://doi.org/10.3390/dna2040020 - 1 Dec 2022
Cited by 9 | Viewed by 3084
Abstract
DNA glycosylases promote genomic stability by initiating base excision repair (BER) in both the nuclear and mitochondrial genomes. Several of these enzymes have overlapping substrate recognition, through which a degree of redundancy in lesion recognition is achieved. For example, OGG1 and NEIL1 both [...] Read more.
DNA glycosylases promote genomic stability by initiating base excision repair (BER) in both the nuclear and mitochondrial genomes. Several of these enzymes have overlapping substrate recognition, through which a degree of redundancy in lesion recognition is achieved. For example, OGG1 and NEIL1 both recognize and release the imidazole-ring-fragmented guanine, FapyGua as part of a common overall pathway to cleanse the genome of damaged bases. However, these glycosylases have many differences, including their differential breadth of substrate specificity, the contrasting chemistries through which base release occurs, the subsequent steps required to complete the BER pathway, and the identity of specific protein-binding partners. Beyond these differences, the complexities and differences of their in vivo biological roles have been primarily elucidated in studies of murine models harboring a knockout of Neil1 or Ogg1, with the diversity of phenotypic manifestations exceeding what might have been anticipated for a DNA glycosylase deficiency. Pathologies associated with deficiencies in nuclear DNA repair include differential cancer susceptibilities, where Ogg1-deficient mice are generally refractory to carcinogenesis, while deficiencies in Neil1-deficient mice confer cancer susceptibility. In contrast to NEIL1, OGG1 functions as a key transcription factor in regulating inflammation and other complex gene cascades. With regard to phenotypes attributed to mitochondrial repair, knockout of either of these genes results in age- and diet-induced metabolic syndrome. The adverse health consequences associated with metabolic syndrome can be largely overcome by expression of a mitochondrial-targeted human OGG1 in both wild-type and Ogg1-deficient mice. The goal of this review is to compare the roles that NEIL1 and OGG1 play in maintaining genomic integrity, with emphasis on insights gained from not only the diverse phenotypes that are manifested in knockout and transgenic mice, but also human disease susceptibility associated with polymorphic variants. Full article
(This article belongs to the Special Issue From Mutation and Repair to Therapeutics)
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15 pages, 1935 KiB  
Article
An Enzyme-Linked Immunosorbent Assay for the Detection of Mitochondrial DNA–Protein Cross-Links from Mammalian Cells
by Wenyan Xu and Linlin Zhao
DNA 2022, 2(4), 264-278; https://doi.org/10.3390/dna2040019 - 11 Nov 2022
Cited by 5 | Viewed by 3045
Abstract
DNA–Protein cross-links (DPCs) are cytotoxic DNA lesions with a protein covalently bound to the DNA. Although much has been learned about the formation, repair, and biological consequences of DPCs in the nucleus, little is known regarding mitochondrial DPCs. This is due in part [...] Read more.
DNA–Protein cross-links (DPCs) are cytotoxic DNA lesions with a protein covalently bound to the DNA. Although much has been learned about the formation, repair, and biological consequences of DPCs in the nucleus, little is known regarding mitochondrial DPCs. This is due in part to the lack of robust and specific methods to measure mitochondrial DPCs. Herein, we reported an enzyme-linked immunosorbent assay (ELISA)-based method for detecting mitochondrial DPCs formed between DNA and mitochondrial transcription factor A (TFAM) in cultured human cells. To optimize the purification and detection workflow, we prepared model TFAM-DPCs via Schiff base chemistry using recombinant human TFAM and a DNA substrate containing an abasic (AP) lesion. We optimized the isolation of TFAM-DPCs using commercial silica gel-based columns to achieve a high recovery yield for DPCs. We evaluated the microplate, DNA-coating solution, and HRP substrate for specific and sensitive detection of TFAM-DPCs. Additionally, we optimized the mtDNA isolation procedure to eliminate almost all nuclear DNA contaminants. For proof of concept, we detected the different levels of TFAM-DPCs in mtDNA from HEK293 cells under different biological conditions. The method is based on commercially available materials and can be amended to detect other types of DPCs in mitochondria. Full article
(This article belongs to the Special Issue From Mutation and Repair to Therapeutics)
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16 pages, 1357 KiB  
Review
The Domino Effect: Nucleosome Dynamics and the Regulation of Base Excision Repair Enzymes
by Julia C. Cook and Sarah Delaney
DNA 2022, 2(4), 248-263; https://doi.org/10.3390/dna2040018 - 10 Nov 2022
Cited by 3 | Viewed by 2874
Abstract
DNA damage is induced by exogenous and endogenous sources, creating a variety of lesions. However, the cellular repair machinery that addresses and corrects this damage must contend with the fact that genomic DNA is sequestered in the nucleoprotein complex of chromatin. As the [...] Read more.
DNA damage is induced by exogenous and endogenous sources, creating a variety of lesions. However, the cellular repair machinery that addresses and corrects this damage must contend with the fact that genomic DNA is sequestered in the nucleoprotein complex of chromatin. As the minimal unit of DNA compaction, the nucleosome core particle (NCP) is a major determinant of repair and poses unique barriers to DNA accessibility. This review outlines how the base excision repair (BER) pathway is modulated by the NCP and describes the structural and dynamic factors that influence the ability of BER enzymes to find and repair damage. Structural characteristics of the NCP such as nucleobase positioning and occupancy will be explored along with factors that impact the dynamic nature of NCPs to increase mobilization of nucleosomal DNA. We will discuss how altering the dynamics of NCPs initiates a domino effect that results in the regulation of BER enzymes. Full article
(This article belongs to the Special Issue From Mutation and Repair to Therapeutics)
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17 pages, 2313 KiB  
Review
Multi-Faceted Roles of ERCC1-XPF Nuclease in Processing Non-B DNA Structures
by Tonia T. Li and Karen M. Vasquez
DNA 2022, 2(4), 231-247; https://doi.org/10.3390/dna2040017 - 11 Oct 2022
Cited by 4 | Viewed by 3028
Abstract
Genetic instability can result from increases in DNA damage and/or alterations in DNA repair proteins and can contribute to disease development. Both exogenous and endogenous sources of DNA damage and/or alterations in DNA structure (e.g., non-B DNA) can impact genome stability. Multiple repair [...] Read more.
Genetic instability can result from increases in DNA damage and/or alterations in DNA repair proteins and can contribute to disease development. Both exogenous and endogenous sources of DNA damage and/or alterations in DNA structure (e.g., non-B DNA) can impact genome stability. Multiple repair mechanisms exist to counteract DNA damage. One key DNA repair protein complex is ERCC1-XPF, a structure-specific endonuclease that participates in a variety of DNA repair processes. ERCC1-XPF is involved in nucleotide excision repair (NER), repair of DNA interstrand crosslinks (ICLs), and DNA double-strand break (DSB) repair via homologous recombination. In addition, ERCC1-XPF contributes to the processing of various alternative (i.e., non-B) DNA structures. This review will focus on the processing of alternative DNA structures by ERCC1-XPF. Full article
(This article belongs to the Special Issue From Mutation and Repair to Therapeutics)
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10 pages, 2283 KiB  
Article
Size- and Stereochemistry-Dependent Transcriptional Bypass of DNA Alkyl Phosphotriester Adducts in Mammalian Cells
by Ying Tan, Jiabin Wu, Garrit Clabaugh, Lin Li, Hua Du and Yinsheng Wang
DNA 2022, 2(4), 221-230; https://doi.org/10.3390/dna2040016 - 5 Oct 2022
Cited by 4 | Viewed by 2366
Abstract
Environmental, endogenous and therapeutic alkylating agents can react with internucleotide phosphate groups in DNA to yield alkyl phosphotriester (PTE) adducts. Alkyl-PTEs are induced at relatively high frequencies and are persistent in mammalian tissues; however, their biological consequences in mammalian cells have not been [...] Read more.
Environmental, endogenous and therapeutic alkylating agents can react with internucleotide phosphate groups in DNA to yield alkyl phosphotriester (PTE) adducts. Alkyl-PTEs are induced at relatively high frequencies and are persistent in mammalian tissues; however, their biological consequences in mammalian cells have not been examined. Herein, we assessed how alkyl-PTEs with different alkyl group sizes and stereochemical configurations (SP and RP diastereomers of Me and nPr) affect the efficiency and fidelity of transcription in mammalian cells. We found that, while the RP diastereomer of Me- and nPr-PTEs constituted moderate and strong blockages to transcription, respectively, the SP diastereomer of the two lesions did not appreciably perturb transcription efficiency. In addition, none of the four alkyl-PTEs induced mutant transcripts. Furthermore, polymerase η assumed an important role in promoting transcription across the SP-Me-PTE, but not any of other three lesions. Loss of other translesion synthesis (TLS) polymerases tested, including Pol κ, Pol ι, Pol ξ and REV1, did not alter the transcription bypass efficiency or mutation frequency for any of the alkyl-PTE lesions. Together, our study provided important new knowledge about the impact of alkyl-PTE lesions on transcription and expanded the substrate pool of Pol η in transcriptional bypass. Full article
(This article belongs to the Special Issue From Mutation and Repair to Therapeutics)
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16 pages, 5162 KiB  
Review
Contributing Factors for Mutagenic DNA Lesion Bypass by DNA Polymerase Eta (polη)
by Hunmin Jung
DNA 2022, 2(4), 205-220; https://doi.org/10.3390/dna2040015 - 28 Sep 2022
Cited by 6 | Viewed by 2726
Abstract
The integrity of DNA replication is under constant threat from various exogenous and endogenous factors along with some epigenetic factors. When there is damage to the genome, cells respond to the damage in two major ways, DNA damage repair and DNA damage tolerance. [...] Read more.
The integrity of DNA replication is under constant threat from various exogenous and endogenous factors along with some epigenetic factors. When there is damage to the genome, cells respond to the damage in two major ways, DNA damage repair and DNA damage tolerance. One of the major mechanisms for DNA damage tolerance is DNA lesion bypass, which is performed by specific DNA polymerases called Y-family DNA polymerases including DNA polymerase eta (polη). Ever since the discovery of polη’s unique role in bypassing cyclobutane pyrimidine dimer (CPD), a wide range of DNA lesions have been experimentally shown to be bypassed by polη. The structural study of polη was greatly boosted by the first elucidation of the N-terminal catalytic domain of polη by X-ray crystallography in 2010. Ever since, a lot of polη catalytic domain crystal structures have been published, which were complexed with an incoming nucleotide and a lesion containing DNA including pyrimidine dimers, cisplatin GpG adduct, 8-oxoguanine (oxoG), 8-oxoadenine (oxoA), N7-methylguanine (N7mG), O6-methylguanine (O6mG), hypoxanthine (HX), and many others. Though polη’s active site is known to be rigid with few conformational changes, there are several contributing factors that could facilitate the lesion bypass such as catalytic metals, syn–anti conformational equilibrium, tautomerization, and specific residues of polη. Each of these components are discussed in detail in this review. Full article
(This article belongs to the Special Issue From Mutation and Repair to Therapeutics)
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13 pages, 3670 KiB  
Article
W Chromosome Evolution by Repeated Recycling in the Frog Glandirana rugosa
by Mitsuaki Ogata, Foyez Shams, Yuri Yoshimura, Tariq Ezaz and Ikuo Miura
DNA 2022, 2(3), 172-184; https://doi.org/10.3390/dna2030012 - 1 Aug 2022
Cited by 4 | Viewed by 3559
Abstract
The Y or W sex chromosome of a heteromorphic pair is usually heterochromatinised and degenerated. However, whether chromosome degeneration constantly proceeds toward an extreme end is not fully understood. Here, we present a case of intermittent evolution of W chromosomes caused by interpopulation [...] Read more.
The Y or W sex chromosome of a heteromorphic pair is usually heterochromatinised and degenerated. However, whether chromosome degeneration constantly proceeds toward an extreme end is not fully understood. Here, we present a case of intermittent evolution of W chromosomes caused by interpopulation hybridisation in the Japanese soil-frog, Glandirana rugosa. This species includes two heteromorphic sex chromosome systems, which are separated into geographic populations, namely the XY and ZW groups. In this study, to uncover the evolutionary mechanisms of the heterogeneous W chromosomes, we genetically investigated the geographic differentiation of the ZW populations along with the closely located XY populations. Analysis of mitochondrial cytochrome b sequences detected three distinct clades, named ZW1, ZW2, and ZW3. High throughput analyses of nuclear genomic DNA showed that autosomal alleles of XY populations were deeply introgressed into the ZW3 sub-group. Based on the genotypes of sex-linked single nucleotide polymorphisms, W-borne androgen receptor gene expression, and WW developmental mortality, we concluded that the X chromosomes were recycled to W chromosomes. Upon inclusion of two cases from another group, Neo-ZW, we observed that the X chromosomes were recycled independently at least four times to the new W chromosomes: a repetition of degeneration and resurrection. Full article
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18 pages, 730 KiB  
Review
Mitochondrial DNA: Consensuses and Controversies
by Inna Shokolenko and Mikhail Alexeyev
DNA 2022, 2(2), 131-148; https://doi.org/10.3390/dna2020010 - 10 Jun 2022
Cited by 8 | Viewed by 9214
Abstract
In the course of its short history, mitochondrial DNA (mtDNA) has made a long journey from obscurity to the forefront of research on major biological processes. mtDNA alterations have been found in all major disease groups, and their significance remains the subject of [...] Read more.
In the course of its short history, mitochondrial DNA (mtDNA) has made a long journey from obscurity to the forefront of research on major biological processes. mtDNA alterations have been found in all major disease groups, and their significance remains the subject of intense research. Despite remarkable progress, our understanding of the major aspects of mtDNA biology, such as its replication, damage, repair, transcription, maintenance, etc., is frustratingly limited. The path to better understanding mtDNA and its role in cells, however, remains torturous and not without errors, which sometimes leave a long trail of controversy behind them. This review aims to provide a brief summary of our current knowledge of mtDNA and highlight some of the controversies that require attention from the mitochondrial research community. Full article
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15 pages, 4704 KiB  
Article
Physlr: Next-Generation Physical Maps
by Amirhossein Afshinfard, Shaun D. Jackman, Johnathan Wong, Lauren Coombe, Justin Chu, Vladimir Nikolic, Gokce Dilek, Yaman Malkoç, René L. Warren and Inanc Birol
DNA 2022, 2(2), 116-130; https://doi.org/10.3390/dna2020009 - 10 Jun 2022
Cited by 4 | Viewed by 3173
Abstract
While conventional physical maps helped build most of the reference genomes we use today, generating the maps was prohibitively expensive, and the technology was abandoned in favor of whole-genome shotgun sequencing (WGS). However, genome assemblies generated using WGS data are often less contiguous. [...] Read more.
While conventional physical maps helped build most of the reference genomes we use today, generating the maps was prohibitively expensive, and the technology was abandoned in favor of whole-genome shotgun sequencing (WGS). However, genome assemblies generated using WGS data are often less contiguous. We introduce Physlr, a tool that leverages long-range information provided by some WGS technologies to construct next-generation physical maps. These maps have many potential applications in genome assembly and analysis, including, but not limited to, scaffolding. In this study, using experimental linked-read datasets from two humans, we used Physlr to construct chromosome-scale physical maps (NGA50s of 52 Mbp and 70 Mbp). We also demonstrated how these physical maps can help scaffold human genome assemblies generated using various sequencing technologies and assembly tools. Across all experiments, Physlr substantially improved the contiguity of baseline assemblies over state-of-the-art linked-read scaffolders. Full article
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12 pages, 1272 KiB  
Article
Updating the Phylogeography and Temporal Evolution of Mitochondrial DNA Haplogroup U8 with Special Mention to the Basques
by Vicente M. Cabrera
DNA 2022, 2(2), 104-115; https://doi.org/10.3390/dna2020008 - 7 Apr 2022
Viewed by 4667
Abstract
Mitochondrial DNA phylogenetic and phylogeographic studies have been very useful in reconstructing the history of modern humans. In addition, recent advances in ancient DNA techniques have enabled direct glimpses of the human past. Taking advantage of these possibilities, I carried out a spatiotemporal [...] Read more.
Mitochondrial DNA phylogenetic and phylogeographic studies have been very useful in reconstructing the history of modern humans. In addition, recent advances in ancient DNA techniques have enabled direct glimpses of the human past. Taking advantage of these possibilities, I carried out a spatiotemporal study of the rare and little-studied mtDNA haplogroup U8. Today, U8, represented by its main branches U8a and U8b, has a wide western Eurasian range but both with average frequencies below 1%. It is known that, in Paleolithic times, U8 reached high frequencies in European hunter-gatherers. However, it is pertinent to precise that only lineages belonging to U8a and U8c, a sister branch of U8b, were detected at that time. In spite of its wide geographic implantation, U8c was extinct after the Last Glacial Maximum, but U8a subsisted until the present day, although it never reached its high Paleolithic frequencies. U8a is detected mainly in northern and western Europe including the Basques, testifying to a minor maternal Paleolithic continuity. In this respect, it is worth mentioning that Basques show more U8-based affinities with continental European than with Mediterranean populations. On the contrary, coalescent ages of the most ancient U8b clades point to a Paleolithic diversification in the Caucasus and the Middle Eastern areas. U8b-derived branches reached eastern Europe since the Mesolithic. Subsequent Neolithic and post-Neolithic expansions widen its ranges in continental Europe and the Mediterranean basin, including northern Africa, albeit always as a minor clade that accompanied other, more representative, mitochondrial lineages. Full article
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18 pages, 4792 KiB  
Article
The Nature and Chromosomal Landscape of Endogenous Retroviruses (ERVs) Integrated in the Sheep Nuclear Genome
by Sarbast Ihsan Mustafa, Trude Schwarzacher and John S. Heslop-Harrison
DNA 2022, 2(1), 86-103; https://doi.org/10.3390/dna2010007 - 16 Mar 2022
Cited by 2 | Viewed by 3342
Abstract
Endogenous retroviruses (ERVs) represent genomic components of retroviral origin that are found integrated in the genomes of various species of vertebrates. These genomic elements have been widely characterized in model organisms and humans. However, composition and abundances of ERVs have not been categorized [...] Read more.
Endogenous retroviruses (ERVs) represent genomic components of retroviral origin that are found integrated in the genomes of various species of vertebrates. These genomic elements have been widely characterized in model organisms and humans. However, composition and abundances of ERVs have not been categorized fully in all domestic animals. The advent of next generation sequencing technologies, development of bioinformatics tools, availability of genomic databases, and molecular cytogenetic techniques have revolutionized the exploration of the genome structure. Here, we investigated the nature, abundance, organization and assembly of ERVs and complete genomes of Jaagsiekte sheep retrovirus (JSRV) from high-throughput sequencing (HTS) data from two Iraqi domestic sheep breeds. We used graph-based read clustering (RepeatExplorer), frequency analysis of short motifs (k-mers), alignment to reference genome assemblies and fluorescent in situ hybridization (FISH). Three classes of ERVs were identified with the total genomic proportions of 0.55% from all analyzed whole genome sequencing raw reads, while FISH to ovine metaphase chromosomes exhibited abundant centromeric to dispersed distribution of these ERVs. Furthermore, the complete genomes of JSRV of two Iraqi sheep breeds were assembled and phylogenetically clustered with the known enJSRV proviruses in sheep worldwide. Characterization of partial and complete sequences of mammalian ERVs is valuable in providing insights into the genome landscape, to help with future genome assemblies, and to identify potential sources of disease when ERVs become active. Full article
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18 pages, 4423 KiB  
Review
Nucleases and Co-Factors in DNA Replication Stress Responses
by Jac A. Nickoloff, Neelam Sharma, Lynn Taylor, Sage J. Allen and Robert Hromas
DNA 2022, 2(1), 68-85; https://doi.org/10.3390/dna2010006 - 1 Mar 2022
Cited by 5 | Viewed by 4868
Abstract
DNA replication stress is a constant threat that cells must manage to proliferate and maintain genome integrity. DNA replication stress responses, a subset of the broader DNA damage response (DDR), operate when the DNA replication machinery (replisome) is blocked or replication forks collapse [...] Read more.
DNA replication stress is a constant threat that cells must manage to proliferate and maintain genome integrity. DNA replication stress responses, a subset of the broader DNA damage response (DDR), operate when the DNA replication machinery (replisome) is blocked or replication forks collapse during S phase. There are many sources of replication stress, such as DNA lesions caused by endogenous and exogenous agents including commonly used cancer therapeutics, and difficult-to-replicate DNA sequences comprising fragile sites, G-quadraplex DNA, hairpins at trinucleotide repeats, and telomeres. Replication stress is also a consequence of conflicts between opposing transcription and replication, and oncogenic stress which dysregulates replication origin firing and fork progression. Cells initially respond to replication stress by protecting blocked replisomes, but if the offending problem (e.g., DNA damage) is not bypassed or resolved in a timely manner, forks may be cleaved by nucleases, inducing a DNA double-strand break (DSB) and providing a means to accurately restart stalled forks via homologous recombination. However, DSBs pose their own risks to genome stability if left unrepaired or misrepaired. Here we focus on replication stress response systems, comprising DDR signaling, fork protection, and fork processing by nucleases that promote fork repair and restart. Replication stress nucleases include MUS81, EEPD1, Metnase, CtIP, MRE11, EXO1, DNA2-BLM, SLX1-SLX4, XPF-ERCC1-SLX4, Artemis, XPG, and FEN1. Replication stress factors are important in cancer etiology as suppressors of genome instability associated with oncogenic mutations, and as potential cancer therapy targets to enhance the efficacy of chemo- and radiotherapeutics. Full article
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12 pages, 883 KiB  
Review
Quality Control of Cell Lines Using DNA as Target
by José Antonio Carrillo-Ávila, Purificación Catalina and Rocío Aguilar-Quesada
DNA 2022, 2(1), 44-55; https://doi.org/10.3390/dna2010004 - 16 Feb 2022
Cited by 3 | Viewed by 4595
Abstract
Cell lines are a widely used pre-clinical models for biomedical research. The accessibility and the relative simplicity of facilities necessary for the use of cell lines, along with the large number of potential applications, encourage many researchers to choose this model. However, the [...] Read more.
Cell lines are a widely used pre-clinical models for biomedical research. The accessibility and the relative simplicity of facilities necessary for the use of cell lines, along with the large number of potential applications, encourage many researchers to choose this model. However, the access to cell lines from a non-confident source or through the interlaboratory exchange results in uncontrollable cell lines of uncertain quality. Furthermore, the possibility of using cell lines as an endless resource through multiple passages can contribute to this uncontrolled scenario, the main consequence of which is the lack of reproducibility between the research results. Different initiatives have emerged to promote the best practices regarding the use of cell lines and minimize the effect on the scientific results reported, including comprehensive quality control in the frame of Good Cell Culture Practice (GCCP). Cell Banks, research infrastructures for the professional distribution of biological material of high and known quality and origin, are committed with these initiatives. Many of the quality controls used to test different attributes of cell lines are based on DNA. This review describes quality control protocols of cell lines whose target molecule is DNA, and details the scope or purpose and their corresponding functionality. Full article
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21 pages, 12848 KiB  
Article
Central Role of C2H2-Type Zinc Finger-Containing Genes in Pediatric Brain Tumors
by Dimitrios S. Kanakoglou, Andromachi Pampalou, Lina S. Malakou, Eleftheria Lakiotaki, Theodoros Loupis, Dimitrios M. Vrachnos, Panayiotis D. Glekas, Alexia Klonou, Angeliki-Ioanna Giannopoulou, Madison Carpenter, Penelope Korkolopoulou and Christina Piperi
DNA 2022, 2(1), 1-21; https://doi.org/10.3390/dna2010001 - 3 Jan 2022
Viewed by 4034
Abstract
Zinc fingers consist of one of the most abundant motifs in transcription factors and DNA-binding proteins. Recent studies provide evidence on the pathological implication of zinc finger proteins in various neurodevelopmental disorders and malignancies but their role in pediatric brain tumors is largely [...] Read more.
Zinc fingers consist of one of the most abundant motifs in transcription factors and DNA-binding proteins. Recent studies provide evidence on the pathological implication of zinc finger proteins in various neurodevelopmental disorders and malignancies but their role in pediatric brain tumors is largely unexplored. To this end, we investigated the differential expression of zinc finger-containing genes along with relevant biological processes and pathways among four main brain tumor categories (pilocytic astrocytomas, ependymomas, medulloblastomas and glioblastomas). By employing an extended bioinformatic toolset, we performed a preliminary in silico study in order to identify the expression of zinc finger-containing genes and associated functions in pediatric brain tumors. Our data analysis reveals the prominent role of C2H2-type zinc finger-containing genes in the molecular mechanisms underlying pediatric brain tumors followed by the Ring and PHD finger types. Significant dysregulation of ABLIM2 and UHFR1 genes was detected in all tumor types drawing attention to the dysregulation of cell polarization process and Ubiquitin-Proteasome System (UPS) in the pathogenesis of pediatric brain tumors. Moreover, significant gene clustering was observed in multiple locations with two highly visible clusters revealing a contrast in gene regulation between medulloblastomas and the other three brain tumor types, indicating a promising area of future research. Full article
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9 pages, 540 KiB  
Article
Incidence, Reproductive Outcome, and Economic Impact of Reciprocal Translocations in the Domestic Pig
by Nicole M. Lewis, Claudia C. Rathje, Carla Canedo-Ribeiro, Lisa M. Bosman, Lucas G. Kiazim, Rebecca L. Jennings, Rebecca E. O’Connor, Giuseppe Silvestri and Darren K. Griffin
DNA 2021, 1(2), 68-76; https://doi.org/10.3390/dna1020007 - 9 Oct 2021
Cited by 2 | Viewed by 3650
Abstract
Pigs (Sus scrofa) have vast economic importance, with pork accounting for over 30% of the global meat consumption. Chromosomal abnormalities, and in particular reciprocal translocations (RTs), are an important cause of hypoprolificacy (litter size reduction) in pigs. However, these do not [...] Read more.
Pigs (Sus scrofa) have vast economic importance, with pork accounting for over 30% of the global meat consumption. Chromosomal abnormalities, and in particular reciprocal translocations (RTs), are an important cause of hypoprolificacy (litter size reduction) in pigs. However, these do not necessarily present with a recognizable phenotype and may cause significant economic losses for breeders when undetected. Here, we present a reappraisal of the incidence of RTs across several European pig herds, using contemporary methodology, as well as an analysis modelling the economic impact of these abnormalities. Molecular cytogenetic investigation was completed by karyotyping and/or multiprobe FISH (fluorescence in situ hybridisation) between 2016–2021, testing 2673 animals. We identified 19 types of chromosome abnormalities, the prevalence of these errors in the database was 9.1%, and the estimated incidence of de novo errors was 0.90%. Financial modelling across different scenarios revealed the potential economic impact of an undetected RT, ranging from £69,802 for an individual affected terminal boar in a commercial farm selling weaned pigs, to £51,215,378 for a genetics company with an undetected RT in a dam line boar used in a nucleus farm. Moreover, the added benefits of screening by FISH instead of karyotyping were estimated, providing a strong case for proactive screening by this approach. Full article
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11 pages, 2510 KiB  
Article
Demethylation of Non-CpG Sites in DNA Is Initiated by TET2 5-Methylcytosine Dioxygenase
by Aninda Sundar Dey, Chayan Bhattacharya, Yihong Guan, Babal Kant Jha and Mridul Mukherji
DNA 2021, 1(1), 26-36; https://doi.org/10.3390/dna1010004 - 21 Sep 2021
Viewed by 4191
Abstract
In the mammalian genome, cytosine methylation predominantly occurs at CpG sites. In addition, a number of recent studies have uncovered extensive C5 cytosine methylation (5mC) at non-CpG (5mCpH, where H = A/C/T) sites. Little is known about the enzyme responsible for active demethylation [...] Read more.
In the mammalian genome, cytosine methylation predominantly occurs at CpG sites. In addition, a number of recent studies have uncovered extensive C5 cytosine methylation (5mC) at non-CpG (5mCpH, where H = A/C/T) sites. Little is known about the enzyme responsible for active demethylation of 5mCpH sites. Using a very sensitive and quantitative LC–MS/MS method, we demonstrate that the human TET2, an iron (II)- and 2OG-dependent dioxygenase, which is a frequently mutated gene in several myeloid malignancies, as well as in a number of other types of cancers, can oxidize 5mCpH sites in double-stranded DNA in vitro. Similar to oxidation of 5mCpG, oxidation of 5mC at CpH sites produces 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxycytosine (5caC) bases in DNA. After 5mCpG, which is the most preferred substrate, TET2 prefers 5mCpC as a substrate, followed by 5mCpA and then 5mCpT. Since the TDG/BER pathway and deformylation or decarboxylation of 5fC or 5caC, respectively, can convert 5fCpH and 5caCpH to an unmodified cytosine base in DNA, our results suggest a novel demethylation pathway of 5mCpH sites initiated by TET2 dioxygenase. Full article
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