Genome Assembly of a Relict Arabian Species of Daphnia O. F. Müller (Crustacea: Cladocera) Adapted to the Desert Life

The water flea Daphnia O.F. Müller 1776 (Crustacea: Cladocera) is an important model of recent evolutionary biology. Here, we report a complete genome of Daphnia (Ctenodaphnia) arabica (Crustacea: Cladocera), recently described species endemic to deserts of the United Arab Emirates. In this study, genome analysis of D. arabica was carried out to investigate its genomic differences, complexity as well as its historical origins within the subgenus Daphnia (Ctenodaphnia). Hybrid genome assembly of D. arabica resulted in ~116 Mb of the assembled genome, with an N50 of ~1.13 Mb (BUSCO score of 99.2%). From the assembled genome, in total protein coding, 5374 tRNA and 643 rRNA genes were annotated. We found that the D. arabica complete genome differed from those of other Daphnia species deposited in the NCBI database but was close to that of D. cf. similoides. However, its divergence time estimate sets D. arabica in the Mesozoic, and our demographic analysis showed a great reduction in its genetic diversity compared to other Daphnia species. Interestingly, the population expansion in its diversity occurred during the megadrought climate around 100 Ka ago, reflecting the adaptive feature of the species to arid and drought-affected environments. Moreover, the PFAM comparative analysis highlights the presence of the important domain SOSS complex subunit C in D. arabica, which is missing in all other studied species of Daphnia. This complex consists of a few subunits (A, B, C) working together to maintain the genome stability (i.e., promoting the reparation of DNA under stress). We propose that this domain could play a role in maintaining the fitness and survival of this species in the desert environment. The present study will pave the way for future research to identify the genes that were gained or lost in this species and identify which of these were key factors to its adaptation to the harsh desert environment.     

Mitochondrial DNA Alterations in Glioblastoma (GBM)

Glioblastoma (GBM) is an extremely aggressive tumor originating from neural stem cells of the central nervous system, which has high histopathological and genomic diversity. Mitochondria are cellular organelles associated with the regulation of cellular metabolism, redox signaling, energy generation, regulation of cell proliferation, and apoptosis. Accumulation of mutations in mitochondrial DNA (mtDNA) leads to mitochondrial dysfunction that plays an important role in GBM pathogenesis, favoring abnormal energy and reactive oxygen species production and resistance to apoptosis and to chemotherapeutic agents. The present review summarizes the known mitochondrial DNA alterations related to GBM, their cellular and metabolic consequences, and their association with diagnosis, prognosis, and treatment.     

Roles of Mitochondrial DNA Damage in Kidney Diseases: A New Biomarker

The kidney is a mitochondria-rich organ, and kidney diseases are recognized as mitochondria-related pathologies. Intact mitochondrial DNA (mtDNA) maintains normal mitochondrial function. Mitochondrial dysfunction caused by mtDNA damage, including impaired mtDNA replication, mtDNA mutation, mtDNA leakage, and mtDNA methylation, is involved in the progression of kidney diseases. Herein, we review the roles of mtDNA damage in different setting of kidney diseases, including acute kidney injury (AKI) and chronic kidney disease (CKD). In a variety of kidney diseases, mtDNA damage is closely associated with loss of kidney function. The level of mtDNA in peripheral serum and urine also reflects the status of kidney injury. Alleviating mtDNA damage can promote the recovery of mitochondrial function by exogenous drug treatment and thus reduce kidney injury. In short, we conclude that mtDNA damage may serve as a novel biomarker for assessing kidney injury in different causes of renal dysfunction, which provides a new theoretical basis for mtDNA-targeted intervention as a therapeutic option for kidney diseases.     

大型溞(Daphnia magna)控藻生态修复技术在典型南亚热带湖泊中的应用

以典型南亚热带富营养化水体——海珠湿地景观水体和荔湾湖为例,采用大型溞摄食藻类、有机悬浮物颗粒迅速提高水体透明度,恢复沉水植物群落,逐步完善修复系统的结构和功能。结果表明:修复后海珠湿地景观水体主要水质指标NH_3–N、TN、TP和CODcr分别在1mg/L、1.5mg/L、0.01mg/L和10mg/L以下,SD在2m左右;荔湾湖修复后水质也基本保持在地表Ⅲ水标准(GB3838–2002)。实践证明,大型溞控藻生态修复技术能够适应南亚热带富营养化水体,效果显著,这也为典型华南富营养化水体的生态修复提供一种有效的治理技术。     

Genes and Pathways Involved in Adult Onset Disorders Featuring Muscle Mitochondrial DNA Instability

Replication and maintenance of mtDNA entirely relies on a set of proteins encoded by the nuclear genome, which include members of the core replicative machinery, proteins involved in the homeostasis of mitochondrial dNTPs pools or deputed to the control of mitochondrial dynamics and morphology. Mutations in their coding genes have been observed in familial and sporadic forms of pediatric and adult-onset clinical phenotypes featuring mtDNA instability. The list of defects involved in these disorders has recently expanded, including mutations in the exo-/endo-nuclease flap-processing proteins MGME1 and DNA2, supporting the notion that an enzymatic DNA repair system actively takes place in mitochondria. The results obtained in the last few years acknowledge the contribution of next-generation sequencing methods in the identification of new disease loci in small groups of patients and even single probands. Although heterogeneous, these genes can be conveniently classified according to the pathway to which they belong. The definition of the molecular and biochemical features of these pathways might be helpful for fundamental knowledge of these disorders, to accelerate genetic diagnosis of patients and the development of rational therapies. In this review, we discuss the molecular findings disclosed in adult patients with muscle pathology hallmarked by mtDNA instability.     

Mitochondrial and Nuclear DNA Damage and Repair in Age-Related Macular Degeneration

Aging and oxidative stress seem to be the most important factors in the pathogenesis of age-related macular degeneration (AMD), a condition affecting many elderly people in the developed world. However, aging is associated with the accumulation of oxidative damage in many biomolecules, including DNA. Furthermore, mitochondria may be especially important in this process because the reactive oxygen species produced in their electron transport chain can damage cellular components. Therefore, the cellular response to DNA damage, expressed mainly through DNA repair, may play an important role in AMD etiology. In several studies the increase in mitochondrial DNA (mtDNA) damage and mutations, and the decrease in the efficacy of DNA repair have been correlated with the occurrence and the stage of AMD. It has also been shown that mitochondrial DNA accumulates more DNA lesions than nuclear DNA in AMD. However, the DNA damage response in mitochondria is executed by nucleus-encoded proteins, and thus mutagenesis in nuclear DNA (nDNA) may affect the ability to respond to mutagenesis in its mitochondrial counterpart. We reported that lymphocytes from AMD patients displayed a higher amount of total endogenous basal and oxidative DNA damage, exhibited a higher sensitivity to hydrogen peroxide and UV radiation, and repaired the lesions induced by these factors less effectively than did cells from control individuals. We postulate that poor efficacy of DNA repair (i.e., is impaired above average for a particular age) when combined with the enhanced sensitivity of retinal pigment epithelium cells to environmental stress factors, contributes to the pathogenesis of AMD. Collectively, these data suggest that the cellular response to both mitochondrial and nuclear DNA damage may play an important role in AMD pathogenesis.     

Retroconversion of Docosapentaenoic Acid (n-6): an Alternative Pathway for Biosynthesis of Arachidonic Acid in Daphnia magna

The aim of this study was to assess metabolic pathways for arachidonic acid (20:4n-6) biosynthesis in Daphnia magna. Neonates of D. magna were maintained on [¹³C] enriched Scenedesmus obliquus and supplemented with liposomes that contained separate treatments of unlabeled docosapentaenoic acid (22:5n-6), 20:4n-6, linoleic acid (18:2n-6) or oleic acid (18:1n-9). Daphnia in the control treatment, without any supplementary fatty acids (FA) containing only trace amounts of 20:4n-6 (~0.3 % of all FA). As expected, the highest proportion of 20:4n-6 (~6.3 %) was detected in Daphnia that received liposomes supplemented with this FA. Higher availability of 18:2n-6 in the diet increased the proportion of 18:2n-6 in Daphnia, but the proportion of 20:4n-6 was not affected. Daphnia supplemented with 22:5n-6 contained ~3.5 % 20:4n-6 in the lipids and FA specific stable isotope analyses validated that the increase in the proportion of 20:4n-6 was due to retroconversion of unlabeled 22:5n-6. These results suggest that chain shortening of 22:5n-6 is a more efficient pathway to synthesize 20:4n-6 in D. magna than elongation and desaturation of 18:2n-6. These results may at least partially explain the discrepancies noticed between phytoplankton FA composition and the expected FA composition in freshwater cladocerans. Finally, retroconversion of dietary 22:5n-6 to 20:4n-6 indicates Daphnia efficiently retain long chain n-6 FA in lake food webs, which might be important for the nutritional ecology of fish.     

Extranuclear Inheritance of Mitochondrial Genome and Epigenetic Reprogrammability of Chromosomal Telomeres in Somatic Cell Cloning of Mammals

The effectiveness of somatic cell nuclear transfer (SCNT) in mammals seems to be still characterized by the disappointingly low rates of cloned embryos, fetuses, and progeny generated. These rates are measured in relation to the numbers of nuclear-transferred oocytes and can vary depending on the technique applied to the reconstruction of enucleated oocytes. The SCNT efficiency is also largely affected by the capability of donor nuclei to be epigenetically reprogrammed in a cytoplasm of reconstructed oocytes. The epigenetic reprogrammability of donor nuclei in SCNT-derived embryos appears to be biased, to a great extent, by the extranuclear (cytoplasmic) inheritance of mitochondrial DNA (mtDNA) fractions originating from donor cells. A high frequency of mtDNA heteroplasmy occurrence can lead to disturbances in the intergenomic crosstalk between mitochondrial and nuclear compartments during the early embryogenesis of SCNT-derived embryos. These disturbances can give rise to incorrect and incomplete epigenetic reprogramming of donor nuclei in mammalian cloned embryos. The dwindling reprogrammability of donor nuclei in the blastomeres of SCNT-derived embryos can also be impacted by impaired epigenetic rearrangements within terminal ends of donor cell-descended chromosomes (i.e., telomeres). Therefore, dysfunctions in epigenetic reprogramming of donor nuclei can contribute to the enhanced attrition of telomeres. This accelerates the processes of epigenomic aging and replicative senescence in the cells forming various tissues and organs of cloned fetuses and progeny. For all the above-mentioned reasons, the current paper aims to overview the state of the art in not only molecular mechanisms underlying intergenomic communication between nuclear and mtDNA molecules in cloned embryos but also intrinsic determinants affecting unfaithful epigenetic reprogrammability of telomeres. The latter is related to their abrasion within somatic cell-inherited chromosomes.     

Mitochondrial DNA Consensus Calling and Quality Filtering for Constructing Ancient Human Mitogenomes: Comparison of Two Widely Applied Methods

Retrieving high-quality endogenous ancient DNA (aDNA) poses several challenges, including low molecular copy number, high rates of fragmentation, damage at read termini, and potential presence of exogenous contaminant DNA. All these factors complicate a reliable reconstruction of consensus aDNA sequences in reads from high-throughput sequencing platforms. Here, we report findings from a thorough evaluation of two alternative tools (ANGSD and schmutzi) aimed at overcoming these issues and constructing high-quality ancient mitogenomes. Raw genomic data (BAM/FASTQ) from a total of 17 previously published whole ancient human genomes ranging from the 14th to the 7th millennium BCE were retrieved and mitochondrial consensus sequences were reconstructed using different quality filters, with their accuracy measured and compared. Moreover, the influence of different sequence parameters (number of reads, sequenced bases, mean coverage, and rate of deamination and contamination) as predictors of derived sequence quality was evaluated. Complete mitogenomes were successfully reconstructed for all ancient samples, and for the majority of them, filtering substantially improved mtDNA consensus calling and haplogroup prediction. Overall, the schmutzi pipeline, which estimates and takes into consideration exogenous contamination, appeared to have the edge over the much faster and user-friendly alternative method (ANGSD) in moderate to high coverage samples (>1,000,000 reads). ANGSD, however, through its read termini trimming filter, showed better capabilities in calling the consensus sequence from low-quality samples. Among all the predictors of overall sample quality examined, the strongest correlation was found for the available number of sequence reads and bases. In the process, we report a previously unassigned haplogroup (U3b) for an Early Chalcolithic individual from Southern Anatolia/Northern Levant.     

A Novel Mitochondrial Genome Fragmentation Pattern in the Buffalo Louse Haematopinus tuberculatus (Psocodea: Haematopinidae)

Sucking lice are obligate ectoparasites of mammalian hosts, causing serious public health problems and economic losses worldwide. It is well known that sucking lice have fragmented mitochondrial (mt) genomes, but many remain undetermined. To better understand patterns of mt genome fragmentation in the sucking lice, we sequenced the mt genome of the buffalo louse Haematopinus tuberculatus using next-generation sequencing (NGS). The mt genome of H. tuberculatus has ten circular minichromosomes containing a total of 37 genes. Each minichromosome is 2.9–5.0 kb long and carries one to eight genes plus one large non-coding region. The number of mt minichromosomes of H. tuberculatus (ten) is different from those of congeneric species (horse louse H. asini, domestic pig louse H. suis and wild pig louse H. apri) and other sucking lice. Two events (gene translocation and merger of mt minichromosome) are observed in Haematopinus. Compared to other studies, our phylogeny generated from mt genome datasets showed a different topology, suggesting that inclusion of data other than mt genomes would be required to resolve phylogeny of sucking lice. To our knowledge, this is the first report of a ten mt minichromosomes genome in sucking lice, which opens a new outlook into unexplored mt genome fragmentation patterns in sucking lice.     

Anatomical Laser Microdissection of the Ileum Reveals mtDNA Depletion Recovery in A Mitochondrial Neuro-Gastrointestinal Encephalomyopathy (MNGIE) Patient Receiving Liver Transplant

mitochondrial neuro-gastrointestinal encephalomyopathy (MNGIE) is a rare genetic disorder characterized by thymidine phosphorylase (TP) enzyme defect. The absence of TP activity induces the imbalance of mitochondrial nucleotide pool, leading to impaired mitochondrial DNA (mtDNA) replication and depletion. Since mtDNA is required to ensure oxidative phosphorylation, metabolically active tissues may not achieve sufficient energy production. The only effective life-saving approach in MNGIE has been the permanent replacement of TP via allogeneic hematopoietic stem cell or liver transplantation. However, the follow-up of transplanted patients showed that gut tissue changes do not revert and fatal complications, such as massive gastrointestinal bleeding, can occur. The purpose of this study was to clarify whether the reintroduction of TP after transplant can recover mtDNA copy number in a normal range. Using laser capture microdissection and droplet-digital-PCR, we assessed the mtDNA copy number in each layer of full-thickness ileal samples of a naive MNGIE cohort vs. controls and in a patient pre- and post-TP replacement. The treatment led to a significant recovery of gut tissue mtDNA amount, thus showing its efficacy. Our results indicate that a timely TP replacement is needed to maximize therapeutic success before irreversible degenerative tissue changes occur in MNGIE.     

Magnesium Isoglycyrrhizinate Reduces the Target-Binding Amount of Cisplatin to Mitochondrial DNA and Renal Injury through SIRT3

Nephrotoxicity is the dose-limiting factor of cisplatin treatment. Magnesium isoglycyrrhizinate (MgIG) has been reported to ameliorate renal ischemia–reperfusion injury. This study aimed to investigate the protective effect and possible mechanisms of MgIG against cisplatin-induced nephrotoxicity from the perspective of cellular pharmacokinetics. We found that cisplatin predominantly accumulated in mitochondria of renal tubular epithelial cells, and the amount of binding with mitochondrial DNA (mtDNA) was more than twice that with nuclear DNA (nDNA). MgIG significantly lowered the accumulation of cisplatin in mitochondria and, in particular, the degree of target-binding to mtDNA. MgIG notably ameliorated cisplatin-induced changes in mitochondrial membrane potential, morphology, function, and cell viability, while the magnesium donor drugs failed to work. In a mouse model, MgIG significantly alleviated cisplatin-caused renal dysfunction, pathological changes of renal tubules, mitochondrial ultrastructure variations, and disturbed energy metabolism. Both in vitro and in vivo data showed that MgIG recovered the reduction of NAD⁺-related substances and NAD⁺-dependent deacetylase sirtuin-3 (SIRT3) level caused by cisplatin. Furthermore, SIRT3 knockdown weakened the protective effect of MgIG on mitochondria, while SIRT3 agonist protected HK-2 cells from cisplatin and specifically reduced platinum-binding activity with mtDNA. In conclusion, MgIG reduces the target-binding amount of platinum to mtDNA and exerts a protective effect on cisplatin-induced renal injury through SIRT3, which may provide a new strategy for the treatment of cisplatin-induced nephrotoxicity.     

De Novo Hybrid Assembly of the Salvia miltiorrhiza Mitochondrial Genome Provides the First Evidence of the Multi-Chromosomal Mitochondrial DNA Structure of Salvia Species

Salvia miltiorrhiza has been an economically important medicinal plant. Previously, an S. miltiorrhiza mitochondrial genome (mitogenome) assembled from Illumina short reads, appearing to be a single circular molecule, has been published. Based on the recent reports on the plant mitogenome structure, we suspected that this conformation does not accurately represent the complexity of the S. miltiorrhiza mitogenome. In the current study, we assembled the mitogenome of S. miltiorrhiza using the PacBio and Illumina sequencing technologies. The primary structure of the mitogenome contained two mitochondrial chromosomes (MC1 and MC2), which corresponded to two major conformations, namely, Mac1 and Mac2, respectively. Using two approaches, including (1) long reads mapping and (2) polymerase chain reaction amplification followed by Sanger sequencing, we observed nine repeats that can mediate recombination. We predicted 55 genes, including 33 mitochondrial protein-coding genes (PCGs), 3 rRNA genes, and 19 tRNA genes. Repeat analysis identified 112 microsatellite repeats and 3 long-tandem repeats. Phylogenetic analysis using the 26 shared PCGs resulted in a tree that was congruent with the phylogeny of Lamiales species in the APG IV system. The analysis of mitochondrial plastid DNA (MTPT) identified 16 MTPTs in the mitogenome. Moreover, the analysis of nucleotide substitution rates in Lamiales showed that the genes atp4, ccmB, ccmFc, and mttB might have been positively selected. The results lay the foundation for future studies on the evolution of the Salvia mitogenome and the molecular breeding of S. miltiorrhiza.     

Experimental and theoretical studies on the photoinduced acute toxicity of a series of anthraquinone derivatives towards the water flea (Daphnia magna)

The photoinduced acute toxicity of fourteen anthraquinone derivatives to Daphnia magna was investigated. Whilst in the dark, all of dyes exhibited no observable toxicity at the maximum test concentration used, in the presence of visible light, all of the colorants, with the exception of three nitroanthraquinones, were acutely toxic towards the water flea over a wide range of medium effective concentrations (4.3–4186.7 nmol L⁻¹). Generally, the acute toxicity of the dyes was higher in the presence of full-spectrum simulated solar radiation compared with visible light; in addition, the photoinduced toxicity of the dyes increased with increase in their hydrophobicity. The energy gap between the lowest unoccupied molecular orbital and the highest occupied molecular orbital was found to indicate the relative photoinduced toxicity of the dyes. Time-dependent density functional theory calculations revealed that singlet oxygen and the superoxide anion could be generated through direct energy transfer or autoionization of the excited state of the dyes.     

Comprehensive Molecular Characterization of the Mitochondrial Genome of the Takin Lungworm Varestrongylus eleguneniensis (Strongylida: Protostrongylidae)

The takin lungworm Varestrongylus eleguneniensis (Strongylida: Protostrongylidae) causes lethal bronchopneumonia and represents severe threats to captive and wild populations. However, until now there has been very limited information available concerning the molecular epidemiology and evolutionary biology of V. eleguneniensis. Mitochondrial genomes (mtDNAs) can provide resources for investigations in these areas and, therefore, can assist with the surveillance and control of this lungworm. Herein, the complete mtDNA of V. eleguneniensis was sequenced and characterized with Illumina pipeline analyses. This circular genome (13,625 bp) encoded twelve protein-coding genes (PCGs), two rRNAs, and twenty-two tRNAs, with notable levels of AT and GC skews. Comparative genomics revealed a purifying selection among PCGs, with cox1 and nad6 having the lowest and the highest evolutionary rate, respectively. Genome-wide phylogenies showed a close relationship between V. eleguneniensis and Protostrongylus rufescens in Strongylida. Single gene (PCGs or rRNAs)-based phylogenies indicated that cox1 and nad5 genes shared the same family-level topology with that inferred from genomic datasets, suggesting that both genes could be suitable genetic markers for evolutionary and phylogenetic studies of Strongylida species. This was the first mtDNA of any member of the genus Varestrongylus, and its comprehensive molecular characterization represents a new resource for systematic, population genetic and evolutionary biological studies of Varestrongylus lungworms in wildlife.