Dataset Information

Composition and Structure of Arabidopsis thaliana Extrachromosomal Circular DNAs Revealed by Nanopore Sequencing.

ABSTRACT: Extrachromosomal circular DNAs (eccDNAs) are enigmatic DNA molecules that have been detected in a range of organisms. In plants, eccDNAs have various genomic origins and may be derived from transposable elements. The structures of individual eccDNA molecules and their dynamics in response to stress are poorly understood. In this study, we showed that nanopore sequencing is a useful tool for the detection and structural analysis of eccDNA molecules. Applying nanopore sequencing to the eccDNA molecules of epigenetically stressed Arabidopsis plants grown under various stress treatments (heat, abscisic acid, and flagellin), we showed that TE-derived eccDNA quantity and structure vary dramatically between individual TEs. Epigenetic stress alone did not cause eccDNA up-regulation, whereas its combination with heat stress triggered the generation of full-length and various truncated eccDNAs of the ONSEN element. We showed that the ratio between full-length and truncated eccDNAs is TE- and condition-dependent. Our work paves the way for further elucidation of the structural features of eccDNAs and their connections with various biological processes, such as eccDNA transcription and eccDNA-mediated TE silencing.

SUBMITTER: Merkulov P

PROVIDER: S-EPMC10255303 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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Composition and Structure of <i>Arabidopsis thaliana</i> Extrachromosomal Circular DNAs Revealed by Nanopore Sequencing.

Merkulov Pavel P Egorova Ekaterina E Kirov Ilya I

Plants (Basel, Switzerland) 20230530 11

Extrachromosomal circular DNAs (eccDNAs) are enigmatic DNA molecules that have been detected in a range of organisms. In plants, eccDNAs have various genomic origins and may be derived from transposable elements. The structures of individual eccDNA molecules and their dynamics in response to stress are poorly understood. In this study, we showed that nanopore sequencing is a useful tool for the detection and structural analysis of eccDNA molecules. Applying nanopore sequencing to the eccDNA mole ...[more]

PMID: 37299157

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Project description:BackgroundEsophageal squamous cell carcinoma (ESCC) is the leading cause of cancer-related mortality. While recent studies have documented the presence of extrachromosomal circular DNAs (eccDNAs) in various tumors, to date, there have been no studies examining the distribution and function of eccDNAs in ESCC.MethodsThe eccDNAs from three surgically matched ESCC tissue samples were extracted and amplified by rolling circle amplification after removal of linear DNA and mitochondrial circular DNA. High-throughput eccDNA sequencing and bioinformatics analysis was performed to study the distribution pattern and the level of eccDNA expression. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed on the genes associated with the differentially expressed eccDNAs. Five up-regulated and five down-regulated candidate eccDNAs were validated by routine polymerase chain reaction (PCR), TOPO-TA cloning and Sanger sequencing. The nucleotides flanking the eccDNA junctions were analyzed to explore the mechanisms of eccDNA formation.ResultsA total of 184,557 eccDNAs was identified. The overall length distribution ranged from 33 to 968,842 base pairs (bp), with the peak at approximately 360 bp. These eccDNAs mainly originated from 5'- and 3'-untranslated regions (UTRs), and rarely from exons, introns, LINE, or Alu repeat regions. The chromosome distribution, length distribution, and genomic annotation of the eccDNAs were comparable between ESCC samples and matched normal epithelium. Nevertheless, 16,031 eccDNAs were found to be differentially expressed between ESCC and matched normal epithelium, including 10,126 up-regulated eccDNAs and 5,905 down-regulated eccDNAs. GO analysis and KEGG pathway analysis showed enriched in cancer pathways, mitogen-activated protein kinase (MAPK) pathway, GTPase-related activity, and cytoskeleton function. PCR, TOPO-TA cloning, and Sanger sequencing validated the junctional sites of five up-regulated candidate eccDNAs and four other unexpected eccDNAs. A repeat nucleotide pattern between the position flanking the start site and that flanking the end site was detected.ConclusionsThis study demonstrated the genome-wide presence of eccDNAs, explored the differential expression of eccDNAs, and revealed the potential mechanisms of eccDNAs in ESCC. This work provides further insights into our understanding of genome plasticity, the role of eccDNAs in ESCC, and may contribute to the development of potential clinical therapies.

Project description:BackgroundThe role of focal amplifications and extrachromosomal circular DNA (eccDNA) is still uncertain in prostate adenocarcinoma (PRAD). Here, we first mapped the global characterizations of eccDNA and then investigate the characterization of eccDNA-amplified key differentially expressed encoded genes (eKDEGs) in the progression, immune response and immunotherapy of PRAD.MethodsCircular_seq was used in conjunction with the TCGA-PRAD transcriptome dataset to sequence, annotate, and filter for eccDNA-amplified differentially expressed coding genes (eDEGs) in PRAD and para-cancerous normal prostate tissues. Afterwards, risk models were created and eKDEGs linked to the PRAD prognosis were identified using Cox and Lasso regression analysis. The immune microenvironment of the risk model was quantified using a variety of immunological algorithms, which also identified its characteristics with regard to immunotherapy, immune response, and immune infiltration.ResultsIn this research, there was no significant difference in the size, type, and chromosomal distribution of eccDNA in PRAD and para-cancerous normal prostate tissues. However, 4,290 differentially expressed eccDNAs were identified and 1,981 coding genes were amplified. Following that, 499 eDEGs were tested in conjunction with the transcriptome dataset from TCGA-PRAD. By using Cox and Lasso regression techniques, ZNF330 and PITPNM3 were identified as eKDEGs of PRAD, and a new PRAD risk model was conducted based on this. Survival analysis showed that the high-risk group of this model was associated with poor prognosis and validated in external data. Immune infiltration analysis showed that the model risks affected immune cell infiltration in PRAD, not only mediating changes in immune cell function, but also correlating with immunophenotyping. Furthermore, the high-risk group was negatively associated with anti-CTLA-4/anti-PD-1 response and mutational burden. In addition, Tumor Immune Dysfunction and Exclusion analyses showed that high-risk group was more prone to immune escape. Drug sensitivity analyses identified 10 drugs, which were instructive for PRAD treatment.ConclusionZNF330 and PITPNM are the eKDEGs for PRAD, which can be used as potential new prognostic markers. The two-factor combined risk model can effectively assess the survival and prognosis of PRAD patients, but also can predict the different responses of immunotherapy to PRAD patients, which may provide new ideas for PRAD immunotherapy.

Dataset Information

Composition and Structure of Arabidopsis thaliana Extrachromosomal Circular DNAs Revealed by Nanopore Sequencing.

Publications

Composition and Structure of <i>Arabidopsis thaliana</i> Extrachromosomal Circular DNAs Revealed by Nanopore Sequencing.

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