Pub Date : 2025-03-01Epub Date: 2025-01-01DOI: 10.3390/dna5010001
Elizabeth Walsh, Tran Zen B Torres, Brian C Prince, Claudia Rückert
Background/objectives: Culex species mosquitoes are globally distributed and transmit several pathogens that impact animal and public health, including West Nile virus, Usutu virus, and Plasmodium relictum. Despite their relevance, Culex species are less widely studied than Aedes and Anopheles mosquitoes. To expand the genetic tools used to study Culex mosquitoes, we previously developed an optimized plasmid for transient Cas9 and single-guide RNA (sgRNA) expression in Culex quinquefasciatus cells to generate gene knockouts. Here, we established a monoclonal cell line that consistently expresses Cas9 and can be used for screens to determine gene function or antiviral activity.
Methods: We used this system to perform the successful gene editing of seven genes and subsequent testing for potential antiviral effects, using a simple single-guide RNA (sgRNA) transfection and subsequent virus infection.
Results: We were able to show antiviral effects for the Cx. quinquefasciatus genes dicer-2, argonaute-2b, vago, piwi5, piwi6a, and cullin4a. In comparison to the RNAi-mediated gene silencing of dicer-2, argonaute-2b, and piwi5, our Cas9/sgRNA approach showed an enhanced ability to detect antiviral effects.
Conclusions: We propose that this cell line offers a new tool for studying gene function in Cx. quinquefasciatus mosquitoes that avoids the use of RNAi. This short study also serves as a proof-of-concept for future gene knock-ins in these cells. Our cell line expands the molecular resources available for vector competence research and will support the design of future research strategies to reduce the transmission of mosquito-borne diseases.
{"title":"Generation of Cas9 Knock-In <i>Culex quinquefasciatus</i> Mosquito Cells.","authors":"Elizabeth Walsh, Tran Zen B Torres, Brian C Prince, Claudia Rückert","doi":"10.3390/dna5010001","DOIUrl":"10.3390/dna5010001","url":null,"abstract":"<p><strong>Background/objectives: </strong><i>Culex</i> species mosquitoes are globally distributed and transmit several pathogens that impact animal and public health, including West Nile virus, Usutu virus, and <i>Plasmodium relictum</i>. Despite their relevance, <i>Culex</i> species are less widely studied than <i>Aedes</i> and <i>Anopheles</i> mosquitoes. To expand the genetic tools used to study <i>Culex</i> mosquitoes, we previously developed an optimized plasmid for transient Cas9 and single-guide RNA (sgRNA) expression in <i>Culex quinquefasciatus</i> cells to generate gene knockouts. Here, we established a monoclonal cell line that consistently expresses Cas9 and can be used for screens to determine gene function or antiviral activity.</p><p><strong>Methods: </strong>We used this system to perform the successful gene editing of seven genes and subsequent testing for potential antiviral effects, using a simple single-guide RNA (sgRNA) transfection and subsequent virus infection.</p><p><strong>Results: </strong>We were able to show antiviral effects for the <i>Cx. quinquefasciatus</i> genes <i>dicer-2</i>, <i>argonaute-2b</i>, <i>vago</i>, <i>piwi5</i>, <i>piwi6a</i>, and <i>cullin4a</i>. In comparison to the RNAi-mediated gene silencing of <i>dicer-2</i>, <i>argonaute-2b</i>, and <i>piwi5</i>, our Cas9/sgRNA approach showed an enhanced ability to detect antiviral effects.</p><p><strong>Conclusions: </strong>We propose that this cell line offers a new tool for studying gene function in <i>Cx. quinquefasciatus</i> mosquitoes that avoids the use of RNAi. This short study also serves as a proof-of-concept for future gene knock-ins in these cells. Our cell line expands the molecular resources available for vector competence research and will support the design of future research strategies to reduce the transmission of mosquito-borne diseases.</p>","PeriodicalId":72835,"journal":{"name":"DNA","volume":"5 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11823230/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143434012","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-09-01Epub Date: 2024-08-01DOI: 10.3390/dna4030015
Kamendra Kumar, Albert J Fornace, Shubhankar Suman
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.
氧化应激介导的生物分子损伤是电离辐射(IR)损伤的一个特征,会导致基因组不稳定和慢性健康影响。具体来说,据报道,在许多组织和生物流体中,红外照射数月后,氧化 DNA 损伤会出现与剂量和线性能量转移(LET)相关的持续性增加。众所周知,与低辐射光子辐射相反,与低辐射红外辐射相比,高辐射红外辐射即使在亚致死剂量下也会导致更高的 DNA 损伤累积。在深空环境(即地球磁层之外)中,高能谱红外辐射非常普遍,暴露在这种辐射下可能会损害宇航员的健康。因此,开发评估和监测 DNA 氧化损伤水平的生物标志物有助于及早发现健康风险,并及时进行干预。在公认的氧化 DNA 损伤生物标志物中,8-氧代-7,8-二氢-2'-脱氧鸟苷(8-OxodG)已成为一种有希望的候选标志物,表明存在慢性氧化应激。据报道,在接受同等剂量的低辐射和高辐射红外照射后,8-OxodG 会表现出不同的水平。本综述讨论了 8-OxodG 作为高强辐射诱导的慢性应激的潜在生物标志物,特别强调了其潜在来源、形成、修复机制和检测方法。此外,这篇综述还探讨了高激光辐射红外照射的病理生物学影响及其与 8-OxodG 的关联。了解高强度线性红外辐射诱导的慢性氧化应激、全身 8-OxodG 水平及其潜在健康风险之间的关联,可为开发全面的健康监测生物标志物系统提供一个框架,以保障宇航员在太空任务期间的健康,并优化长期健康结果。
{"title":"8-OxodG: A Potential Biomarker for Chronic Oxidative Stress Induced by High-LET Radiation.","authors":"Kamendra Kumar, Albert J Fornace, Shubhankar Suman","doi":"10.3390/dna4030015","DOIUrl":"https://doi.org/10.3390/dna4030015","url":null,"abstract":"<p><p>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.</p>","PeriodicalId":72835,"journal":{"name":"DNA","volume":"4 3","pages":"221-238"},"PeriodicalIF":0.0,"publicationDate":"2024-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11391509/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142302463","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The three 31 nucleotide minihelix tRNA evolution theorem describes the evolution of type I and type II tRNAs to the last nucleotide. In databases, type I and type II tRNA V loops (V for variable) were improperly aligned, but alignment based on the theorem is accurate. Type II tRNA V arms were a 3′-acceptor stem (initially CCGCCGC) ligated to a 5′-acceptor stem (initially GCGGCGG). The type II V arm evolved to form a stem–loop–stem. In Archaea, tRNALeu and tRNASer are type II. In Bacteria, tRNALeu, tRNASer, and tRNATyr are type II. The trajectory of the type II V arm is determined by the number of unpaired bases just 5′ of the Levitt base (Vmax). For Archaea, tRNALeu has two unpaired bases, and tRNASer has one unpaired base. For Bacteria, tRNATyr has two unpaired bases, tRNALeu has one unpaired base, and tRNASer has zero unpaired bases. Thus, the number of synonymous type II tRNA sets is limited by the possible trajectory set points of the arm. From the analysis of aminoacyl-tRNA synthetase structures, contacts to type II V arms appear to adjust allosteric tension communicated primarily via tRNA to aminoacylating and editing active sites. To enhance allostery, it appears that type II V arm end loop contacts may tend to evolve to V arm stem contacts.
三个 31 核苷酸小螺旋 tRNA 演化定理描述了 I 型和 II 型 tRNA 演化到最后一个核苷酸的过程。在数据库中,I型和II型tRNA的V环(V代表变量)排列不当,但基于该定理的排列是准确的。II 型 tRNA V 臂是一个 3′受体茎(最初为 CCGCCGC)连接到一个 5′受体茎(最初为 GCGGCGG)。II 型 V 臂进化成茎-环-茎。在古生菌中,tRNALeu 和 tRNASer 属于 II 型。在细菌中,tRNALeu、tRNASer 和 tRNATyr 都是 II 型。II 型 V 臂的轨迹由莱维特碱基 5′ 处的未配对碱基数量(Vmax)决定。对于古菌,tRNALeu 有两个未配对碱基,tRNASer 有一个未配对碱基。细菌的 tRNATyr 有两个未配对碱基,tRNALeu 有一个未配对碱基,而 tRNASer 的未配对碱基为零。因此,同义的第二类 tRNA 组的数量受到臂的可能轨迹集点的限制。从对氨基酰-tRNA 合成酶结构的分析来看,与 II 型 V 臂的接触似乎主要是通过 tRNA 与氨基酰化和编辑活性位点的沟通来调整异构张力。为了增强异构张力,II 型 V 臂末端环接触似乎倾向于演变为 V 臂茎接触。
{"title":"Origin of Type II tRNA Variable Loops, Aminoacyl-tRNA Synthetase Allostery from Distal Determinants, and Diversification of Life","authors":"Lei Lei, Zachary Frome Burton","doi":"10.3390/dna4030017","DOIUrl":"https://doi.org/10.3390/dna4030017","url":null,"abstract":"The three 31 nucleotide minihelix tRNA evolution theorem describes the evolution of type I and type II tRNAs to the last nucleotide. In databases, type I and type II tRNA V loops (V for variable) were improperly aligned, but alignment based on the theorem is accurate. Type II tRNA V arms were a 3′-acceptor stem (initially CCGCCGC) ligated to a 5′-acceptor stem (initially GCGGCGG). The type II V arm evolved to form a stem–loop–stem. In Archaea, tRNALeu and tRNASer are type II. In Bacteria, tRNALeu, tRNASer, and tRNATyr are type II. The trajectory of the type II V arm is determined by the number of unpaired bases just 5′ of the Levitt base (Vmax). For Archaea, tRNALeu has two unpaired bases, and tRNASer has one unpaired base. For Bacteria, tRNATyr has two unpaired bases, tRNALeu has one unpaired base, and tRNASer has zero unpaired bases. Thus, the number of synonymous type II tRNA sets is limited by the possible trajectory set points of the arm. From the analysis of aminoacyl-tRNA synthetase structures, contacts to type II V arms appear to adjust allosteric tension communicated primarily via tRNA to aminoacylating and editing active sites. To enhance allostery, it appears that type II V arm end loop contacts may tend to evolve to V arm stem contacts.","PeriodicalId":72835,"journal":{"name":"DNA","volume":"9 12","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141921730","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Tamás Cseppentő, Norbert G. Valis, G. Bárány, Bálint Megadja, Attila Heinrich, Nóra M. Magonyi
In forensic DNA laboratories, it is important to conduct internal validations of the commercially available kits of short tandem repeat (STR) loci and to investigate their individual and combined effectiveness. This study aims to report on a comparative investigation of the forensic kits used in our laboratory and their combinations in analysing low-copy-number (LCN) human DNA samples. We used five partly overlapping multiplex kits with different marker configurations from different manufacturers: the NGM SelectTM PCR Amplification Kit, NGM DetectTM, the GlobalFilerTM Amplification Kit (Applied BiosystemTM, Foster City, CA, USA), the PowerPlex® Fusion 6C System (Promega Co., Madison, WI, USA) and the Investigator® 24plex QS Kit (Qiagen GmbH, Hilden, Germany). The efficacy of the kits was scrutinised by specific criteria, such as allelic dropout rate, the individually calculated Likelihood Ratio (LR) of consensus profiles and the LR value of the composite profile produced by the combined profiles of two kits. According to the results, the pairing of PowerPlex® Fusion 6C System and Investigator® 24plex QS produced the lowest, while the pairing of the NGM DetectTM and GlobalFilerTM kits provided the highest LR value. In summary, our study is meant to aid the selection of the optimal kit combination for samples of different qualities.
在法医 DNA 实验室中,对市场上销售的短串联重复(STR)位点试剂盒进行内部验证,并研究其单独和组合的有效性非常重要。本研究旨在报告我们实验室在分析低拷贝数(LCN)人类 DNA 样本时使用的法医试剂盒及其组合的比较调查。我们使用了五种部分重叠的多重试剂盒,它们具有不同的标记配置,分别来自不同的生产商:NGM SelectTM PCR扩增试剂盒、NGM DetectTM、GlobalFilerTM扩增试剂盒(Applied BiosystemTM,美国加利福尼亚州福斯特市)、PowerPlex® Fusion 6C系统(Promega Co.,美国威斯康星州麦迪逊)和Investigator® 24plex QS试剂盒(Qiagen GmbH,德国希尔登)。试剂盒的功效通过特定的标准进行检验,如等位基因丢失率、单独计算的共识图谱的似然比(LR)以及由两个试剂盒的组合图谱产生的复合图谱的 LR 值。结果显示,PowerPlex® Fusion 6C 系统和 Investigator® 24plex QS 配对的 LR 值最低,而 NGM DetectTM 和 GlobalFilerTM 试剂盒配对的 LR 值最高。总之,我们的研究有助于为不同质量的样本选择最佳的试剂盒组合。
{"title":"Comparative Analysis of Five Forensic PCR Kits in Duplets","authors":"Tamás Cseppentő, Norbert G. Valis, G. Bárány, Bálint Megadja, Attila Heinrich, Nóra M. Magonyi","doi":"10.3390/dna4030014","DOIUrl":"https://doi.org/10.3390/dna4030014","url":null,"abstract":"In forensic DNA laboratories, it is important to conduct internal validations of the commercially available kits of short tandem repeat (STR) loci and to investigate their individual and combined effectiveness. This study aims to report on a comparative investigation of the forensic kits used in our laboratory and their combinations in analysing low-copy-number (LCN) human DNA samples. We used five partly overlapping multiplex kits with different marker configurations from different manufacturers: the NGM SelectTM PCR Amplification Kit, NGM DetectTM, the GlobalFilerTM Amplification Kit (Applied BiosystemTM, Foster City, CA, USA), the PowerPlex® Fusion 6C System (Promega Co., Madison, WI, USA) and the Investigator® 24plex QS Kit (Qiagen GmbH, Hilden, Germany). The efficacy of the kits was scrutinised by specific criteria, such as allelic dropout rate, the individually calculated Likelihood Ratio (LR) of consensus profiles and the LR value of the composite profile produced by the combined profiles of two kits. According to the results, the pairing of PowerPlex® Fusion 6C System and Investigator® 24plex QS produced the lowest, while the pairing of the NGM DetectTM and GlobalFilerTM kits provided the highest LR value. In summary, our study is meant to aid the selection of the optimal kit combination for samples of different qualities.","PeriodicalId":72835,"journal":{"name":"DNA","volume":"78 7","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141655580","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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.
{"title":"Efficient Elimination of mtDNA from Mammalian Cells with 2′,3′-Dideoxycytidine","authors":"N. Kozhukhar, Mikhail F. Alexeyev","doi":"10.3390/dna4030013","DOIUrl":"https://doi.org/10.3390/dna4030013","url":null,"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 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.","PeriodicalId":72835,"journal":{"name":"DNA","volume":" 31","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-07-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141677781","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
T. Bianco-Miotto, Sadia Hossain, Nahal Habibi, Dandara G. Haag, J. Grieger
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.
{"title":"Child Telomere Length at 11–12 Years of Age Is Not Associated with Pregnancy Complications","authors":"T. Bianco-Miotto, Sadia Hossain, Nahal Habibi, Dandara G. Haag, J. Grieger","doi":"10.3390/dna4020011","DOIUrl":"https://doi.org/10.3390/dna4020011","url":null,"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 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.","PeriodicalId":72835,"journal":{"name":"DNA","volume":"3 10","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-06-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141356306","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-01Epub Date: 2024-03-30DOI: 10.3390/dna4020006
Nelson C Lau, Vanessa M Macias
Mosquitoes, like Drosophila, are dipterans, the order of "true flies" characterized by a single set of two wings. Drosophila are prime model organisms for biomedical research, while mosquito researchers struggle to establish robust molecular biology in these that are arguably the most dangerous vectors of human pathogens. Both insects utilize the RNA interference (RNAi) pathway to generate small RNAs to silence transposons and viruses, yet details are emerging that several RNAi features are unique to each insect family, such as how culicine mosquitoes have evolved extreme genomic feature differences connected to their unique RNAi features. A major technical difference in the molecular genetic studies of these insects is that generating stable transgenic animals are routine in Drosophila but still variable in stability in mosquitoes, despite genomic DNA-editing advances. By comparing and contrasting the differences in the RNAi pathways of Drosophila and mosquitoes, in this review we propose a hypothesis that transgene DNAs are possibly more intensely targeted by mosquito RNAi pathways and chromatin regulatory pathways than in Drosophila. We review the latest findings on mosquito RNAi pathways, which are still much less well understood than in Drosophila, and we speculate that deeper study into how mosquitoes modulate transposons and viruses with Piwi-interacting RNAs (piRNAs) will yield clues to improving transgene DNA expression stability in transgenic mosquitoes.
{"title":"Transposon and Transgene Tribulations in Mosquitoes: A Perspective of piRNA Proportions.","authors":"Nelson C Lau, Vanessa M Macias","doi":"10.3390/dna4020006","DOIUrl":"10.3390/dna4020006","url":null,"abstract":"<p><p>Mosquitoes, like <i>Drosophila</i>, are dipterans, the order of \"true flies\" characterized by a single set of two wings. <i>Drosophila</i> are prime model organisms for biomedical research, while mosquito researchers struggle to establish robust molecular biology in these that are arguably the most dangerous vectors of human pathogens. Both insects utilize the RNA interference (RNAi) pathway to generate small RNAs to silence transposons and viruses, yet details are emerging that several RNAi features are unique to each insect family, such as how culicine mosquitoes have evolved extreme genomic feature differences connected to their unique RNAi features. A major technical difference in the molecular genetic studies of these insects is that generating stable transgenic animals are routine in <i>Drosophila</i> but still variable in stability in mosquitoes, despite genomic DNA-editing advances. By comparing and contrasting the differences in the RNAi pathways of <i>Drosophila</i> and mosquitoes, in this review we propose a hypothesis that transgene DNAs are possibly more intensely targeted by mosquito RNAi pathways and chromatin regulatory pathways than in <i>Drosophila</i>. We review the latest findings on mosquito RNAi pathways, which are still much less well understood than in <i>Drosophila</i>, and we speculate that deeper study into how mosquitoes modulate transposons and viruses with Piwi-interacting RNAs (piRNAs) will yield clues to improving transgene DNA expression stability in transgenic mosquitoes.</p>","PeriodicalId":72835,"journal":{"name":"DNA","volume":"4 2","pages":"104-128"},"PeriodicalIF":0.0,"publicationDate":"2024-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11286205/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141794184","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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.
电离辐射会诱发多种不同类型的 DNA 损伤。但其特点之一是产生复杂的 DNA 损伤,其中串联 DNA 损伤因其独特的生物特性而备受关注。组织炎症引起的氧化应激以及产生自由基的金属催化反应也会形成这些 DNA 损伤。在本小视图中,我们总结了串联病变的形成以及在特定位点合成后对其进行的复制和修复研究。其中许多病变对传统的碱基切除修复具有抗性,因此只能通过核苷酸切除修复途径进行修复。它们还会阻碍 DNA 复制,当发生病变旁路时,可能会出现严重的错误。其中一些串联 DNA 病变可能会导致衰老、神经系统疾病和癌症。
{"title":"Mutagenesis and Repair of γ-Radiation- and Radical-Induced Tandem DNA Lesions","authors":"A. Basu, Laureen C. Colis, J. H. T. Bacurio","doi":"10.3390/dna4020009","DOIUrl":"https://doi.org/10.3390/dna4020009","url":null,"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 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.","PeriodicalId":72835,"journal":{"name":"DNA","volume":"3 9","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-05-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141006273","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Repetitive DNA sequences are abundant in the human genome and can adopt alternative (i.e., non-B) DNA structures. These sequences contribute to diverse biological functions, including genomic instability. Previously, we found that Z-DNA-, H-DNA- and cruciform DNA-forming sequences are mutagenic, implicating them in cancer etiology. These sequences can stimulate the formation of DNA double-strand breaks (DSBs), causing deletions via cleavage by the endonuclease ERCC1-XPF. Interestingly, the activity of ERCC1-XPF in H-DNA-induced mutagenesis is nucleotide excision repair (NER)-dependent, but its role in Z-DNA-induced mutagenesis is NER-independent. Instead, Z-DNA is processed by ERCC1-XPF in a mechanism dependent on the mismatch repair (MMR) complex, MSH2-MSH3. These observations indicate distinct mechanisms of non-B-induced genomic instability. However, the roles of NER and MMR proteins, as well as additional nucleases (CtIP and MRE11), in the processing of cruciform DNA remain unknown. Here, we present data on the processing of cruciform-forming short inverted repeats (IRs) by DNA repair proteins using mammalian cell-based systems. From this pilot study, we show that, in contrast to H-DNA and Z-DNA, short IRs are processed in a NER- and MMR-independent manner, and the nucleases CtIP and MRE11 suppress short IR-induced genomic instability in mammalian cells.
重复 DNA 序列在人类基因组中非常丰富,可采用替代性(即非 B 型)DNA 结构。这些序列具有多种生物功能,包括基因组不稳定性。此前,我们发现 Z-DNA、H-DNA 和十字形 DNA 形成序列具有致突变性,与癌症病因有关。这些序列可刺激 DNA 双链断裂(DSB)的形成,通过内切酶 ERCC1-XPF 的裂解造成缺失。有趣的是,ERCC1-XPF 在 H-DNA 诱导突变中的活性依赖核苷酸切除修复(NER),但在 Z-DNA 诱导突变中的作用却不依赖 NER。相反,ERCC1-XPF处理Z-DNA的机制依赖于错配修复(MMR)复合物MSH2-MSH3。这些观察结果表明了非 B 诱导的基因组不稳定性的不同机制。然而,NER 和 MMR 蛋白以及其他核酸酶(CtIP 和 MRE11)在十字形 DNA 处理过程中的作用仍然未知。在这里,我们利用基于哺乳动物细胞的系统,展示了 DNA 修复蛋白处理十字形短倒位重复序列(IR)的数据。通过这项试验性研究,我们发现与 H-DNA 和 Z-DNA 不同,短 IR 是以不依赖于 NER 和 MMR 的方式处理的,而核酸酶 CtIP 和 MRE11 能抑制哺乳动物细胞中短 IR 诱导的基因组不稳定性。
{"title":"Exploring the Roles of Different DNA Repair Proteins in Short Inverted Repeat Mediated Genomic Instability: A Pilot Study","authors":"Pooja Mandke, Karen M. Vasquez","doi":"10.3390/dna4020008","DOIUrl":"https://doi.org/10.3390/dna4020008","url":null,"abstract":"Repetitive DNA sequences are abundant in the human genome and can adopt alternative (i.e., non-B) DNA structures. These sequences contribute to diverse biological functions, including genomic instability. Previously, we found that Z-DNA-, H-DNA- and cruciform DNA-forming sequences are mutagenic, implicating them in cancer etiology. These sequences can stimulate the formation of DNA double-strand breaks (DSBs), causing deletions via cleavage by the endonuclease ERCC1-XPF. Interestingly, the activity of ERCC1-XPF in H-DNA-induced mutagenesis is nucleotide excision repair (NER)-dependent, but its role in Z-DNA-induced mutagenesis is NER-independent. Instead, Z-DNA is processed by ERCC1-XPF in a mechanism dependent on the mismatch repair (MMR) complex, MSH2-MSH3. These observations indicate distinct mechanisms of non-B-induced genomic instability. However, the roles of NER and MMR proteins, as well as additional nucleases (CtIP and MRE11), in the processing of cruciform DNA remain unknown. Here, we present data on the processing of cruciform-forming short inverted repeats (IRs) by DNA repair proteins using mammalian cell-based systems. From this pilot study, we show that, in contrast to H-DNA and Z-DNA, short IRs are processed in a NER- and MMR-independent manner, and the nucleases CtIP and MRE11 suppress short IR-induced genomic instability in mammalian cells.","PeriodicalId":72835,"journal":{"name":"DNA","volume":"20 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-04-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140738358","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Since the discovery of transposable elements (TEs) in maize in the 1940s by Barbara McClintock transposable elements have been described as junk, as selfish elements with no benefit to the host, and more recently as major determinants of genome structure and genome evolution. TEs are DNA sequences that are capable of moving to new sites in the genome and making additional copies of themselves while doing so. To limit the propagation of TEs, host silencing mechanisms are directed at transposon-encoded genes that are required for mobilization. The mutagenic properties of TEs, the potential of TEs to form new genes and affect gene expression, together with the host silencing mechanisms, shape eukaryotic genomes and drive genome evolution. While TEs constitute more than half of the genome in many higher eukaryotes, transposable elements in the nematode C. elegans form a relatively small proportion of the genome (approximately 15%). Genetic studies of transposon silencing, and the discovery of RNA interference (RNAi) in C. elegans, propelled Caenorhabditis elegans (C. elegans) to the forefront of studies of RNA-based mechanisms that silence TEs. Here, I will review the transposable elements that are present and active in the C. elegans genome, and the host defense mechanisms that silence these elements.
自 20 世纪 40 年代芭芭拉-麦克林托克(Barbara McClintock)在玉米中发现可转座元件(Transposable elements,TEs)以来,可转座元件一直被描述为垃圾元件、对宿主无益的自私元件,最近则被描述为基因组结构和基因组进化的主要决定因素。转座元件是一种 DNA 序列,能够在基因组中移动到新的位点,并在移动的同时复制自己。为了限制 TE 的传播,宿主沉默机制针对的是转座子编码的基因,这些基因是转座子移动所必需的。转座因子的诱变特性、转座因子形成新基因和影响基因表达的潜力以及宿主沉默机制共同塑造了真核生物基因组,并推动了基因组进化。在许多高等真核生物中,TE 占基因组的一半以上,而在线虫秀丽隐杆线虫中,转座元件只占基因组相对较小的比例(约 15%)。对转座子沉默的遗传学研究以及 RNA 干扰(RNAi)在秀丽隐杆线虫中的发现,将秀丽隐杆线虫(C. elegans)推向了基于 RNA 的 TE 沉默机制研究的前沿。在这里,我将回顾一下在秀丽隐杆线虫基因组中存在和活跃的转座元件,以及使这些元件沉默的宿主防御机制。
{"title":"Activity and Silencing of Transposable Elements in C. elegans","authors":"Sylvia E. J. Fischer","doi":"10.3390/dna4020007","DOIUrl":"https://doi.org/10.3390/dna4020007","url":null,"abstract":"Since the discovery of transposable elements (TEs) in maize in the 1940s by Barbara McClintock transposable elements have been described as junk, as selfish elements with no benefit to the host, and more recently as major determinants of genome structure and genome evolution. TEs are DNA sequences that are capable of moving to new sites in the genome and making additional copies of themselves while doing so. To limit the propagation of TEs, host silencing mechanisms are directed at transposon-encoded genes that are required for mobilization. The mutagenic properties of TEs, the potential of TEs to form new genes and affect gene expression, together with the host silencing mechanisms, shape eukaryotic genomes and drive genome evolution. While TEs constitute more than half of the genome in many higher eukaryotes, transposable elements in the nematode C. elegans form a relatively small proportion of the genome (approximately 15%). Genetic studies of transposon silencing, and the discovery of RNA interference (RNAi) in C. elegans, propelled Caenorhabditis elegans (C. elegans) to the forefront of studies of RNA-based mechanisms that silence TEs. Here, I will review the transposable elements that are present and active in the C. elegans genome, and the host defense mechanisms that silence these elements.","PeriodicalId":72835,"journal":{"name":"DNA","volume":"203 ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140751207","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}