RESUMO
BACKGROUND: Mitochondrial (mt) heteroplasmy can cause adverse biological consequences when deleterious mtDNA mutations accumulate disrupting "normal" mt-driven processes and cellular functions. To investigate the heteroplasmy of such mtDNA changes, we developed a moderate throughput mt isolation procedure to quantify the mt single-nucleotide variant (SNV) landscape in individual mouse neurons and astrocytes. In this study, we amplified mt-genomes from 1645 single mitochondria isolated from mouse single astrocytes and neurons to (1) determine the distribution and proportion of mt-SNVs as well as mutation pattern in specific target regions across the mt-genome, (2) assess differences in mtDNA SNVs between neurons and astrocytes, and (3) study co-segregation of variants in the mouse mtDNA. RESULTS: (1) The data show that specific sites of the mt-genome are permissive to SNV presentation while others appear to be under stringent purifying selection. Nested hierarchical analysis at the levels of mitochondrion, cell, and mouse reveals distinct patterns of inter- and intra-cellular variation for mt-SNVs at different sites. (2) Further, differences in the SNV incidence were observed between mouse neurons and astrocytes for two mt-SNV 9027:G > A and 9419:C > T showing variation in the mutational propensity between these cell types. Purifying selection was observed in neurons as shown by the Ka/Ks statistic, suggesting that neurons are under stronger evolutionary constraint as compared to astrocytes. (3) Intriguingly, these data show strong linkage between the SNV sites at nucleotide positions 9027 and 9461. CONCLUSIONS: This study suggests that segregation as well as clonal expansion of mt-SNVs is specific to individual genomic loci, which is important foundational data in understanding of heteroplasmy and disease thresholds for mutation of pathogenic variants.
Assuntos
Astrócitos , Mutação , Neurônios , Animais , Astrócitos/metabolismo , Camundongos , Neurônios/metabolismo , Heteroplasmia/genética , DNA Mitocondrial/genética , Mitocôndrias/genética , Análise de Sequência de DNA/métodosRESUMO
Somatic cells accumulate genomic alterations with age; however, our understanding of mitochondrial DNA (mtDNA) mosaicism remains limited. Here we investigated the genomes of 2,096 clones derived from three cell types across 31 donors, identifying 6,451 mtDNA variants with heteroplasmy levels of â³0.3%. While the majority of these variants were unique to individual clones, suggesting stochastic acquisition with age, 409 variants (6%) were shared across multiple embryonic lineages, indicating their origin from heteroplasmy in fertilized eggs. The mutational spectrum exhibited replication-strand bias, implicating mtDNA replication as a major mutational process. We evaluated the mtDNA mutation rate (5.0 × 10-8 per base pair) and a turnover frequency of 10-20 per year, which are fundamental components shaping the landscape of mtDNA mosaicism over a lifetime. The expansion of mtDNA-truncating mutations toward homoplasmy was substantially suppressed. Our findings provide comprehensive insights into the origins, dynamics and functional consequences of mtDNA mosaicism in human somatic cells.
Assuntos
DNA Mitocondrial , Mosaicismo , Mutação , Humanos , DNA Mitocondrial/genética , Heteroplasmia/genética , Taxa de Mutação , Mitocôndrias/genética , Genoma Mitocondrial , Replicação do DNA/genética , Feminino , MasculinoRESUMO
Mitochondrial function is critical to continued cellular vitality and is an important contributor to a growing number of human diseases. Mitochondrial dysfunction is typically heterogeneous, mediated through the clonal expansion of mitochondrial DNA (mtDNA) variants in a subset of cells in a given tissue. To date, our understanding of the dynamics of clonal expansion of mtDNA variants has been technically limited to the single cell-level. Here, we report the use of nanobiopsy for subcellular sampling from human tissues, combined with next-generation sequencing to assess subcellular mtDNA mutation load in human tissue from mitochondrial disease patients. The ability to map mitochondrial mutation loads within individual cells of diseased tissue samples will further our understanding of mitochondrial genetic diseases.
Assuntos
DNA Mitocondrial , Heteroplasmia , Sequenciamento de Nucleotídeos em Larga Escala , Mutação , Humanos , DNA Mitocondrial/genética , Heteroplasmia/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Doenças Mitocondriais/genética , Doenças Mitocondriais/patologia , Doenças Mitocondriais/metabolismo , Mitocôndrias/genética , Mitocôndrias/metabolismo , Mitocôndrias/patologiaRESUMO
Plant cells harbor two membrane-bound organelles containing their own genetic material-plastids and mitochondria. Although the two organelles coexist and coevolve within the same plant cells, they differ in genome copy number, intracellular organization, and mode of segregation. How these attributes affect the time to fixation or, conversely, loss of neutral alleles is currently unresolved. Here, we show that mitochondria and plastids share the same mutation rate, yet plastid alleles remain in a heteroplasmic state significantly longer compared with mitochondrial alleles. By analyzing genetic variants across populations of the marine flowering plant Zostera marina and simulating organelle allele dynamics, we examine the determinants of allele segregation and allele fixation. Our results suggest that the bottlenecks on the cell population, e.g. during branching or seeding, and stratification of the meristematic tissue are important determinants of mitochondrial allele dynamics. Furthermore, we suggest that the prolonged plastid allele dynamics are due to a yet unknown active plastid partition mechanism. The dissimilarity between plastid and mitochondrial novel allele fixation at different levels of organization may manifest in differences in adaptation processes. Our study uncovers fundamental principles of organelle population genetics that are essential for further investigations of long-term evolution and molecular dating of divergence events.
Assuntos
Heteroplasmia , Mitocôndrias , Taxa de Mutação , Plastídeos , Plastídeos/genética , Mitocôndrias/genética , Mitocôndrias/metabolismo , AlelosRESUMO
Multicopied mitogenome are prone to mutation during replication often resulting in heteroplasmy. The derived variants in a cell, organ, or an individual animal constitute a mitogene pool. The individual mitogene pool is initiated by a small fraction of the egg mitogene pool. However, the characteristics and relationship between them has not yet been investigated. This study quantitatively analyzed the heteroplasmy landscape, genetic loads, and selection strength of the mitogene pool of egg and hatchling in the silver carp (Hypophthalmichthys molitrix) using high-throughput resequencing. The results showed heteroplasmic sites distribute across the whole mitogenome in both eggs and hatchlings. The dominant substitution was Transversion in eggs and Transition in hatching accounting for 95.23%±2.07% and 85.38%±6.94% of total HP sites, respectively. The total genetic loads were 0.293±0.044 in eggs and 0.228±0.022 in hatchlings (P=0.048). The dN/dS ratio was 58.03±38.98 for eggs and 9.44±3.93 for hatchlings (P=0.037). These results suggest that the mitogenomes were under strong positive selection in eggs with tolerance to variants with deleterious effects, while the selection was positive but much weaker in hatchlings showing marked quality control. Based on these findings, we proposed a trans-generation dynamics model to explain differential development mode of the two mitogene pool between oocyte maturation and ontogenesis of offspring. This study sheds light on significance of mitogene pool for persistence of populations and subsequent integration in ecological studies and conservation practices.
Assuntos
Carpas , Genoma Mitocondrial , Animais , Carpas/genética , Heteroplasmia , Seleção Genética , Sequenciamento de Nucleotídeos em Larga EscalaRESUMO
Molluscan mitochondrial genomes are unusual because they show wide variation in size, radical genome rearrangements and frequently show high variation (> 10%) within species. As progress in understanding this variation has been limited, we used whole genome sequencing of a six-generation matriline of the terrestrial snail Cepaea nemoralis, as well as whole genome sequences from wild-collected C. nemoralis, the sister species C. hortensis, and multiple other snail species to explore the origins of mitochondrial DNA (mtDNA) variation. The main finding is that a high rate of SNP heteroplasmy in somatic tissue was negatively correlated with mtDNA copy number in both Cepaea species. In individuals with under ten mtDNA copies per nuclear genome, more than 10% of all positions were heteroplasmic, with evidence for transmission of this heteroplasmy through the germline. Further analyses showed evidence for purifying selection acting on non-synonymous mutations, even at low frequency of the rare allele, especially in cytochrome oxidase subunit 1 and cytochrome b. The mtDNA of some individuals of Cepaea nemoralis contained a length heteroplasmy, including up to 12 direct repeat copies of tRNA-Val, with 24 copies in another snail, Candidula rugosiuscula, and repeats of tRNA-Thr in C. hortensis. These repeats likely arise due to error prone replication but are not correlated with mitochondrial copy number in C. nemoralis. Overall, the findings provide key insights into mechanisms of replication, mutation and evolution in molluscan mtDNA, and so will inform wider studies on the biology and evolution of mtDNA across animal phyla.
Assuntos
Variações do Número de Cópias de DNA , DNA Mitocondrial , Genoma Mitocondrial , Heteroplasmia , Mutação , Seleção Genética , Caramujos , Animais , Caramujos/genética , DNA Mitocondrial/genética , Heteroplasmia/genética , Polimorfismo de Nucleotídeo ÚnicoRESUMO
BACKGROUND: Mitochondrial heteroplasmy reflects genetic diversity within individuals due to the presence of varying mitochondrial DNA (mtDNA) sequences, possibly affecting mitochondrial function and energy production in cells. Rapid growth during early childhood is a critical development with long-term implications for health and well-being. In this study, we investigated if cord blood mtDNA heteroplasmy is associated with rapid growth at 6 and 12 months and overweight in childhood at 4-6 years. METHODS: This study included 200 mother-child pairs of the ENVIRONAGE birth cohort. Whole mitochondrial genome sequencing was performed to determine mtDNA heteroplasmy levels (in variant allele frequency; VAF) in cord blood. Rapid growth was defined for each child as the difference between WHO-SD scores of predicted weight at either 6 or 12 months and birth weight. Logistic regression models were used to determine the association of mitochondrial heteroplasmy with rapid growth and childhood overweight. Determinants of relevant cord blood mitochondrial heteroplasmies were identified using multiple linear regression models. RESULTS: One % increase in VAF of cord blood MT-D-Loop16362T > C heteroplasmy was associated with rapid growth at 6 months (OR = 1.03; 95% CI: 1.01-1.05; p = 0.001) and 12 months (OR = 1.02; 95% CI: 1.00-1.03; p = 0.02). Furthermore, this variant was associated with childhood overweight at 4-6 years (OR = 1.01; 95% CI 1.00-1.02; p = 0.05). Additionally, rapid growth at 6 months (OR = 3.00; 95% CI: 1.49-6.14; p = 0.002) and 12 months (OR = 4.05; 95% CI: 2.06-8.49; p < 0.001) was also associated with childhood overweight at 4-6 years. Furthermore, we identified maternal age, pre-pregnancy BMI, maternal education, parity, and gestational age as determinants of cord blood MT-D-Loop16362T > C heteroplasmy. CONCLUSIONS: Our findings, based on mitochondrial DNA genotyping, offer insights into the molecular machinery leading to rapid growth in early life, potentially explaining a working mechanism of the development toward childhood overweight.
Assuntos
DNA Mitocondrial , Heteroplasmia , Humanos , Feminino , DNA Mitocondrial/genética , Masculino , Recém-Nascido , Lactente , Pré-Escolar , Heteroplasmia/genética , Sangue Fetal/química , Obesidade Infantil/genética , Criança , Mitocôndrias/genética , Sobrepeso/genética , AdultoRESUMO
The extent of cell-to-cell variation in tumor mitochondrial DNA (mtDNA) copy number and genotype, and the phenotypic and evolutionary consequences of such variation, are poorly characterized. Here we use amplification-free single-cell whole-genome sequencing (Direct Library Prep (DLP+)) to simultaneously assay mtDNA copy number and nuclear DNA (nuDNA) in 72,275 single cells derived from immortalized cell lines, patient-derived xenografts and primary human tumors. Cells typically contained thousands of mtDNA copies, but variation in mtDNA copy number was extensive and strongly associated with cell size. Pervasive whole-genome doubling events in nuDNA associated with stoichiometrically balanced adaptations in mtDNA copy number, implying that mtDNA-to-nuDNA ratio, rather than mtDNA copy number itself, mediated downstream phenotypes. Finally, multimodal analysis of DLP+ and single-cell RNA sequencing identified both somatic loss-of-function and germline noncoding variants in mtDNA linked to heteroplasmy-dependent changes in mtDNA copy number and mitochondrial transcription, revealing phenotypic adaptations to disrupted nuclear/mitochondrial balance.
Assuntos
Núcleo Celular , Variações do Número de Cópias de DNA , DNA Mitocondrial , Genoma Mitocondrial , Neoplasias , Análise de Célula Única , Humanos , DNA Mitocondrial/genética , Análise de Célula Única/métodos , Variações do Número de Cópias de DNA/genética , Núcleo Celular/genética , Neoplasias/genética , Neoplasias/patologia , Linhagem Celular Tumoral , Animais , Mitocôndrias/genética , Sequenciamento Completo do Genoma/métodos , Camundongos , Heteroplasmia/genéticaRESUMO
Mitochondrial DNA (mtDNA) exhibits distinct characteristics distinguishing it from the nuclear genome, necessitating specific analytical methods in genetic studies. This comprehensive review explores the complex role of mtDNA in a variety of genetic studies, including genome-wide, epigenome-wide, and phenome-wide association studies, with a focus on its implications for human traits and diseases. Here, we discuss the structure and gene-encoding properties of mtDNA, along with the influence of environmental factors and epigenetic modifications on its function and variability. Particularly significant are the challenges posed by mtDNA's high mutation rate, heteroplasmy, and copy number variations, and their impact on disease susceptibility and population genetic analyses. The review also highlights recent advances in methodological approaches that enhance our understanding of mtDNA associations, advocating for refined genetic research techniques that accommodate its complexities. By providing a comprehensive overview of the intricacies of mtDNA, this paper underscores the need for an integrated approach to genetic studies that considers the unique properties of mitochondrial genetics. Our findings aim to inform future research and encourage the development of innovative methodologies to better interpret the broad implications of mtDNA in human health and disease.
Assuntos
DNA Mitocondrial , Humanos , DNA Mitocondrial/genética , Variações do Número de Cópias de DNA , Epigênese Genética , Estudo de Associação Genômica Ampla/métodos , Heteroplasmia/genética , Mitocôndrias/genética , Predisposição Genética para DoençaRESUMO
Heteroplasmy occurs when wild-type and mutant mitochondrial DNA (mtDNA) molecules co-exist in single cells1. Heteroplasmy levels change dynamically in development, disease and ageing2,3, but it is unclear whether these shifts are caused by selection or drift, and whether they occur at the level of cells or intracellularly. Here we investigate heteroplasmy dynamics in dividing cells by combining precise mtDNA base editing (DdCBE)4 with a new method, SCI-LITE (single-cell combinatorial indexing leveraged to interrogate targeted expression), which tracks single-cell heteroplasmy with ultra-high throughput. We engineered cells to have synonymous or nonsynonymous complex I mtDNA mutations and found that cell populations in standard culture conditions purge nonsynonymous mtDNA variants, whereas synonymous variants are maintained. This suggests that selection dominates over simple drift in shaping population heteroplasmy. We simultaneously tracked single-cell mtDNA heteroplasmy and ancestry, and found that, although the population heteroplasmy shifts, the heteroplasmy of individual cell lineages remains stable, arguing that selection acts at the level of cell fitness in dividing cells. Using these insights, we show that we can force cells to accumulate high levels of truncating complex I mtDNA heteroplasmy by placing them in environments where loss of biochemical complex I activity has been reported to benefit cell fitness. We conclude that in dividing cells, a given nonsynonymous mtDNA heteroplasmy can be harmful, neutral or even beneficial to cell fitness, but that the 'sign' of the effect is wholly dependent on the environment.
Assuntos
Divisão Celular , Linhagem da Célula , DNA Mitocondrial , Aptidão Genética , Heteroplasmia , Seleção Genética , Análise de Célula Única , Animais , Feminino , Humanos , Camundongos , Divisão Celular/genética , Linhagem Celular , Linhagem da Célula/genética , DNA Mitocondrial/genética , Edição de Genes , Heteroplasmia/genética , Mitocôndrias/genética , Mutação , Análise de Célula Única/métodosRESUMO
T cells have been shown to maintain a lower percentage (heteroplasmy) of the pathogenic m.3243A>G variant (MT-TL1, associated with maternally inherited diabetes and deafness [MIDD] and mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes [MELAS]). The mechanism(s) underlying this purifying selection, however, remain unknown. Here we report that purified patient memory CD4+ T cells have lower bulk m.3243A>G heteroplasmy compared to naïve CD4+ T cells. In vitro activation of naïve CD4+ m.3243A>G patient T cells results in lower bulk m.3243A>G heteroplasmy after proliferation. Finally, m.3243A>G patient T cell receptor repertoire sequencing reveals relative oligoclonality compared to controls. These data support a role for T cell activation in peripheral, purifying selection against high m.3243A>G heteroplasmy T cells at the level of the cell, in a likely cell-autonomous fashion.
Assuntos
Ativação Linfocitária , Síndrome MELAS , Humanos , Síndrome MELAS/genética , Linfócitos T CD4-Positivos/imunologia , Heteroplasmia/genética , RNA de Transferência de Leucina/genética , Masculino , Feminino , DNA Mitocondrial/genética , AdultoRESUMO
Due to the exclusive maternal transmission, oocyte mitochondrial dysfunction reduces fertility rates, affects embryonic development, and programs offspring to metabolic diseases. However, mitochondrial DNA (mtDNA) are vulnerable to mutations during oocyte maturation, leading to mitochondrial nucleotide variations (mtSNVs) within a single oocyte, referring to mtDNA heteroplasmy. Obesity (OB) accounts for more than 40% of women at the reproductive age in the USA, but little is known about impacts of OB on mtSNVs in mature oocytes. It is found that OB reduces mtDNA content and increases mtSNVs in mature oocytes, which impairs mitochondrial energetic functions and oocyte quality. In mature oocytes, OB suppresses AMPK activity, aligned with an increased binding affinity of the ATF5-POLG protein complex to mutated mtDNA D-loop and protein-coding regions. Similarly, AMPK knockout increases the binding affinity of ATF5-POLG proteins to mutated mtDNA, leading to the replication of heteroplasmic mtDNA and impairing oocyte quality. Consistently, AMPK activation blocks the detrimental impacts of OB by preventing ATF5-POLG protein recruitment, improving oocyte maturation and mitochondrial energetics. Overall, the data uncover key features of AMPK activation in suppressing mtSNVs, and improving mitochondrial biogenesis and oocyte maturation in obese females.
Assuntos
Proteínas Quinases Ativadas por AMP , DNA Mitocondrial , Obesidade , Oócitos , Oócitos/metabolismo , Obesidade/metabolismo , Obesidade/genética , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Feminino , Camundongos , Animais , Proteínas Quinases Ativadas por AMP/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Heteroplasmia/genética , Fatores Ativadores da Transcrição/metabolismo , Fatores Ativadores da Transcrição/genética , DNA Polimerase gama/genética , DNA Polimerase gama/metabolismo , Humanos , Mitocôndrias/metabolismo , Mitocôndrias/genéticaRESUMO
Heteroplasmy, the presence of multiple mitochondrial DNA (mtDNA) haplotypes within cells of an individual, is caused by mutation or paternal leakage. However, heteroplasmy is usually resolved to homoplasmy within a few generations because of germ-line bottlenecks; therefore, instances of heteroplasmy are limited in nature. Here, we report heteroplasmy in the ricefish species Oryzias matanensis, endemic to Lake Matano, an ancient lake in Sulawesi Island, in which one individual was known to have many heterozygous sites in the mitochondrial NADH dehydrogenase subunit 2 (ND2) gene. In this study, we cloned the ND2 gene for some additional individuals with heterozygous sites and demonstrated that they are truly heteroplasmic. Phylogenetic analysis revealed that the extra haplotype within the heteroplasmic O. matanensis individuals clustered with haplotypes of O. marmoratus, a congeneric species inhabiting adjacent lakes. This indicated that the heteroplasmy originated from paternal leakage due to interspecific hybridization. The extra haplotype was unique and contained two non-synonymous substitutions. These findings demonstrate that this hybridization-driven heteroplasmy was maintained across generations for a long time to the extent that the extra mitochondria evolved within the new host.
Assuntos
Heteroplasmia , Oryzias , Humanos , Animais , Lagos , Filogenia , Oryzias/genética , DNA Mitocondrial/genéticaRESUMO
Maternally inherited diabetes and deafness (MIDD) is a rare mitochondrial disorder primarily resulting from m.3243A>G mutation. The clinical characteristics of MIDD exhibit significant heterogeneity. Our study aims to delineate these characteristics and determine the potential correlation with m.3243A>G heteroplasmy levels. This retrospective, descriptive study encompassed patients with confirmed m.3243A>G mutation and diabetes mellitus at Seoul National University Hospital. Our cohort comprises 40 patients with MIDD, with a mean age at study enrollment of 33.3±12.9 years and an average % of heteroplasmy of 30.0%± 14.6% in the peripheral blood. The most prevalent comorbidity was hearing loss (90%), followed by albuminuria (61%), seizure (38%), and stroke (33%). We observed a significant negative correlation between % of heteroplasmy and age at diabetes diagnosis. These clinical features can aid in the suspicion of MIDD and further consideration of genetic testing for m.3243A>G mutation.
Assuntos
DNA Mitocondrial , Surdez , Doenças Mitocondriais , Mutação , Humanos , Feminino , Masculino , DNA Mitocondrial/genética , Estudos Retrospectivos , Adulto , República da Coreia/epidemiologia , Doenças Mitocondriais/genética , Doenças Mitocondriais/diagnóstico , Doenças Mitocondriais/epidemiologia , Pessoa de Meia-Idade , Surdez/genética , Adulto Jovem , Diabetes Mellitus/genética , Diabetes Mellitus/epidemiologia , Adolescente , Heteroplasmia , Perda Auditiva/genética , Diabetes Mellitus Tipo 2RESUMO
BACKGROUND: Human mitochondrial heteroplasmy is an extensively investigated phenomenon in the context of medical diagnostics, forensic identification and molecular evolution. However, technical limitations of high-throughput sequencing hinder reliable determination of point heteroplasmies (PHPs) with minor allele frequencies (MAFs) within the noise threshold. RESULTS: To investigate the PHP landscape at an MAF threshold down to 0.1%, we sequenced whole mitochondrial genomes at approximately 7.700x coverage, in multiple technical and biological replicates of longitudinal blood and buccal swab samples from 11 human donors (159 libraries in total). The results obtained by two independent sequencing platforms and bioinformatics pipelines indicate distinctive PHP patterns below and above the 1% MAF cut-off. We found a high inter-individual prevalence of low-level PHPs (MAF < 1%) at polymorphic positions of the mitochondrial DNA control region (CR), their tissue preference, and a tissue-specific minor allele linkage. We also established the position-dependent potential of minor allele expansion in PHPs, and short-term PHP instability in a mitotically active tissue. We demonstrate that the increase in sensitivity of PHP detection to minor allele frequencies below 1% within a robust experimental and analytical pipeline, provides new information with potential applicative value. CONCLUSIONS: Our findings reliably show different mutational loads between tissues at sub-1% allele frequencies, which may serve as an informative medical biomarker of time-dependent, tissue-specific mutational burden, or help discriminate forensically relevant tissues in a single person, close maternal relatives or unrelated individuals of similar phylogenetic background.
Assuntos
Heteroplasmia , Mitocôndrias , Humanos , Filogenia , Mitocôndrias/genética , Sequenciamento de Nucleotídeos em Larga Escala , DNA Mitocondrial/genéticaRESUMO
Massively parallel sequencing (MPS) technology has become the gold standard in mitochondrial DNA research due to its high sensitivity in detecting mtDNA heteroplasmy, a prognostic marker in various medical applications. Various MPS technologies and platforms used for mtDNA analysis exist. Obtaining reliable and sensitive results requires deep and uniform coverage of the entire mtDNA sequence, which is heavily influenced by the choice of library preparation method and sequencing platform. Here, we present a comparison of the sequencing coverage and the ability to heteroplasmy detection using two library preparation protocols (Nextera XT DNA Library Preparation Kit and Nextera DNA Flex Library Preparation Kit) and two different (MiSeq FGx and ISeq 100) Illumina MPS platforms. Our study indicates that the Nextera DNA Flex Library protocol provides a more balanced coverage along the mitogenome and a reliable heteroplasmy detection with both MiSeq and iSeq Illumina MPS systems.
Assuntos
DNA Mitocondrial , Biblioteca Gênica , Heteroplasmia , Sequenciamento de Nucleotídeos em Larga Escala , DNA Mitocondrial/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Heteroplasmia/genética , Análise de Sequência de DNA/métodos , Genoma Mitocondrial/genéticaRESUMO
Biodiversity plays a pivotal role in sustaining ecosystem processes, encompassing diverse biological species, genetic types and the intricacies of ecosystem composition. However, the precise definition of biodiversity at the individual level remains a challenging endeavour. Hill numbers, derived from Rényi's entropy, have emerged as a popular measure of diversity, with a recent unified framework extending their application across various levels, from genetics to ecosystems. In this study, we employ a computational approach to exploring the diversity of mitochondrial heteroplasmy using real-world data. By adopting Hill numbers with q = 2, we demonstrate the feasibility of quantifying mitochondrial heteroplasmy diversity within and between individuals and populations. Furthermore, we investigate the alpha diversity of mitochondrial heteroplasmy among different species, revealing heterogeneity at multiple levels, including mitogenome components and protein-coding genes (PCGs). Our analysis explores large-scale mitochondrial heteroplasmy data in humans, examining the relationship between alpha diversity at the mitogenome components and PCGs level. Notably, we do not find a significant correlation between these two levels. Additionally, we observe significant correlations in alpha diversity between mothers and children in blood samples, exceeding the reported R2 value for allele frequency correlations. Moreover, our investigation of beta diversity and local overlay similarity demonstrates that heteroplasmy variant distributions in different tissues of children more closely resemble those of their mothers. Through systematic quantification and analysis of mitochondrial heteroplasmy diversity, this study enhances our understanding of heterogeneity at multiple levels, from individuals to populations, providing new insights into this fundamental dimension of biodiversity.
Assuntos
Ecossistema , Heteroplasmia , Criança , Humanos , Mitocôndrias/genética , Biodiversidade , DNA Mitocondrial/genéticaRESUMO
BACKGROUND: In recent years, the mitochondria/immune system interaction has been proposed, so that variants of mitochondrial genome and levels of heteroplasmy might deregulate important metabolic processes in fighting infections, such as leprosy. METHODS: We sequenced the whole mitochondrial genome to investigate variants and heteroplasmy levels, considering patients with different clinical forms of leprosy and household contacts. After sequencing, a specific pipeline was used for preparation and bioinformatics analysis to select heteroplasmic variants. RESULTS: We found 116 variants in at least two of the subtypes of the case group (Borderline Tuberculoid, Borderline Lepromatous, Lepromatous), suggesting a possible clinical significance to these variants. Notably, 15 variants were exclusively found in these three clinical forms, of which five variants stand out for being missense (m.3791T > C in MT-ND1, m.5317C > A in MT-ND2, m.8545G > A in MT-ATP8, m.9044T > C in MT-ATP6 and m.15837T > C in MT-CYB). In addition, we found 26 variants shared only by leprosy poles, of which two are characterized as missense (m.4248T > C in MT-ND1 and m.8027G > A in MT-CO2). CONCLUSION: We found a significant number of variants and heteroplasmy levels in the leprosy patients from our cohort, as well as six genes that may influence leprosy susceptibility, suggesting for the first time that the mitogenome might be involved with the leprosy process, distinction of clinical forms and severity. Thus, future studies are needed to help understand the genetic consequences of these variants.
Assuntos
Genoma Mitocondrial , Hanseníase , Humanos , Heteroplasmia , Genoma Mitocondrial/genética , Hanseníase/genética , Mitocôndrias/genéticaRESUMO
The human mitochondrial genome (mtDNA) is a circular DNA molecule with a length of 16.6 kb, which contains a total of 37 genes. Somatic mtDNA mutations accumulate with age and environmental exposure, and some types of mtDNA variants may play a role in carcinogenesis. Recent studies observed mtDNA variants not only in kidney tumors but also in adjacent kidney tissues, and mtDNA dysfunction results in kidney injury, including chronic kidney disease (CKD). To investigate whether a relationship exists between heteroplasmic mtDNA variants and kidney function, we performed ultra-deep sequencing (30,000×) based on long-range PCR of DNA from 77 non-tumor kidney tissues of kidney cancer patients with CKD (stages G1 to G5). In total, this analysis detected 697 single-nucleotide variants (SNVs) and 504 indels as heteroplasmic (0.5% ≤ variant allele frequency (VAF) < 95%), and the total number of detected SNVs/indels did not differ between CKD stages. However, the number of deleterious low-level heteroplasmic variants (pathogenic missense, nonsense, frameshift and tRNA) significantly increased with CKD progression (p < 0.01). In addition, mtDNA copy numbers (mtDNA-CNs) decreased with CKD progression (p < 0.001). This study demonstrates that mtDNA damage, which affects mitochondrial genes, may be involved in reductions in mitochondrial mass and associated with CKD progression and kidney dysfunction.