Project description:We report mitochondrial genome (mtDNA) sequences in purified mouse muscle stem cells at different ages. This study identifies changes in the mitochondrial genome of muscle stem cells during aging.
Project description:Mitochondria have an independent genome (mtDNA) and protein synthesis machinery that coordinately activate for mitochondrial generation. Here, we report that Krebs cycle intermediate, fumarate, connects metabolism to mitobiogenesis through binding to malic enzyme 2 (ME2). Proteomic profiling of ME2-interacting protein reveals that ME2 binds to mitoribosome proteins to regulate mitoribosome assembly and mtDNA-encoded protein production. ME2 also interacts with and modulates deoxyuridine 5-triphosphate nucleotidohydrolase activity to regulate thymidine generation and mtDNA abundance
Project description:Background: Cell free DNA (cfDNA) in plasma has received increasing attention and has been studied in a broad range of clinical conditions implicating inflammation, cancer, and aging. However, few studies have focused on mitochondrial DNA (mtDNA) in the cell free form. This study characterized the size distribution and sequence characteristics of plasma cell free mtDNA (cf mtDNA) in humans.Methods and Results: We optimized DNA isolation and next-generation sequencing library preparation protocols to better retain short DNA fragments from plasma, and applied these optimized methods to plasma samples from patients with sepsis. After massive parallel sequencing, we verified that our methods can retain substantially shorter DNA fragments than the standard isolation method, resulting in an average of 11.5 fold increase in short DNA fragments yield (DNA < 100bp). We report that cf mtDNA in plasma is highly enriched in short-size cfDNA (30 ~ 60 bp), which is much shorter than the value previously reported (~140 bp). Motivated by this unique size distribution, we size-selected short cfDNA fragments from the sequencing library, which further increased the mtDNA recovery rate by an average of 10.4 fold. Using this approach we detected mixtures of different mtDNA sequences, termed heteroplasmy, in plasma from 3 patients. In one patient who previously received bone marrow transplantation, different minor allele frequencies were observed between plasma and white blood cells (WBC) at heteroplasmic mtDNA sites, consistent with mixed-tissue origin for plasma DNA.Conclusion: mtDNA in plasma exists as very short fragments that exhibit mtDNA heteroplasmy distribution differences from that found in a single organ/tissue. This study is the first report of genome wide identification of mtDNA heteroplasmy in human plasma. Our optimized method can be used to investigate the potential utility of cf mtDNA fragments and heteroplasmy as biomarkers in various diseases.
Project description:More than 2x10E9 sequences made on Illumina platform derived from the genome of E14 embryonic stem cells cultured in our laboratory were used to build a database of about 2.7x10E6 single nucleotide variant. The database was validated using other two sequencing datasets from other laboratory and high overlap was observed. The identified variant are enriched on intergenic regions, but several thousands reside on gene exons and regulatory regions, such as promoters, enhancers, splicing site and untranslated regions of RNA, thus indicating high probability of an important functional impact on the molecular biology of this cells. We created a new E14 genome assembly including the new identified variants and used it to map reads from next generation sequencing data generated in our laboratory or in others on E14 cell line. We observed an increase in the number of mapped reads of about 5%. CpG dinucleotide showed the higher variation frequency, probably because of it could be target of DNA methylation. We performed a reduced representation bisulfite sequencing on E14 cell line to test our new genome assembly with respect to the mm9 genome reference. After mapping and methylation status calling, we obtained an increase of about 120,000 called CpG and we avoided about 20,000 wrong CpG calling. genotyping of E14 embryonic stem cells (ESCs) and Reduced representation Bisulfite Sequencing (RRBS) of E14 ESCs.
Project description:Mitochondrial DNA (mtDNA) damage is considered as a possible primary cause of Parkinson’s disease (PD). To explore the issue, mtDNA sequences from whole blood were analyzed in PD patients and controls using a resequencing chip and allelic substitutions were estimated for each nucleotide position (np) along the entire mtDNA sequence. Overall, 58 np showed a different allelic distribution in the two groups; of these, 81% showed an increase of non-reference alleles in PD patients, similar to findings reported in patients with Alzheimer’s disease, albeit in reduced proportion. These results suggest that age-related neurodegenerative diseases could share a mechanism involving mtDNA.
Project description:<p> The casuarina moth (Lymantria xylina) is a notorious forestry pest, posing severe ecological and economic threats due to its destructive defoliation outbreaks and high invasive potential. Despite its significance, a high-quality reference genome has been lacking, limiting molecular-level investigations into its biology and hindering the development of effective pest management strategies. In this study, we report the first chromosome-level genome assembly of L. xylina generated through a combination of illumina short-reads, Oxford Nanopore long-reads, and Hi-C scaffolding. The final assembly spans 977.74 Mb, with 95.17% anchored to 31 pseudo-chromosomes, achieving a scaffold N50 of 34.15 Mb. Importantly, telomeric sequences were identified at both ends of all 31 pseudo-chromosomes, underscoring the exceptional quality and completeness of this reference genome. Quality assessment further revealed a BUSCO completeness of 94.5% and a consensus QV of 31.72. We also annotated 18,484 protein-coding genes, 95.21% of which were functionally assigned, and characterized genome-wide repetitive elements (77.18%).</p><p> Beyond the genome assembly, we generated comprehensive RNA-seq and metabolomic datasets across multiple diapause stages, enabling insights into gene expression dynamics and metabolic regulation during egg development. Together, these resources provide a valuable foundation for studying the genetic basis of host adaptation, invasiveness, and interactions with natural enemies such as nucleopolyhedrovirus and Beauveria bassiana.</p>
Project description:Centromeres are chromosomal regions that serve as platforms for kinetochore assembly and spindle attachments, ensuring accurate chromosome segregation during cell division. Despite functional conservation, centromeric sequences are diverse and usually repetitive across species, making them challenging to assemble and identify. Here, we describe centromeres in the model oomycete Phytophthora sojae by combining long-read sequencing-based genome assembly and chromatin immunoprecipitation for the centromeric histone CENP-A followed by high-throughput sequencing (ChIP-seq). P. sojae centromeres cluster at a single focus in the nucleus at different life stages and during nuclear division. We report a highly contiguous genome assembly of the P. sojae reference strain, which enabled identification of 15 highly enriched CENP-A binding regions as putative centromeres. By focusing on 10 intact regions, we demonstrate that centromeres in P. sojae are regional, spanning 211 to 356 kb. Most of these regions are transposon-rich, poorly transcribed, and lack the euchromatin mark H3K4me2 but are embedded within regions with the heterochromatin marks H3K9me3 and H3K27me3.