Detection and quantification of mitochondrial DNA deletions from next-generation sequence data

Bosworth, Colleen M.; Grandhi, Sneha; Gould, Meetha P.; LaFramboise, Thomas

doi:10.1186/s12859-017-1821-7

Cited by 31 publications

(19 citation statements)

References 27 publications

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“…The functional effect of the variants were predicted using open-source algorithms including PolyPhen-2 131 , PANTHER 132 , Envision 133 , MutationAssessor 134 , MutPred2 135 and SNPs&GO 136 . The mtDNA deletion was also determined via MitoDel 137 and eKLIPse 138 . MitoDel is a tool for detecting and quantifying mtDNA deletions even at low heteroplasmy levels via the BLAT split read mapping method.…”

Section: Methodsmentioning

confidence: 99%

Mitochondrial DNA alterations may influence the cisplatin responsiveness of oral squamous cell carcinoma

Aminuddin

Leong

et al. 2020

Sci Rep

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Cisplatin is the first-line chemotherapeutic agent for the treatment of oral squamous cell carcinoma (OSCC). However, the intrinsic or acquired resistance against cisplatin remains a major obstacle to treatment efficacy in OSCC. Recently, mitochondrial DNA (mtDNA) alterations have been reported in a variety of cancers. However, the role of mtDNA alterations in OSCC has not been comprehensively studied. In this study, we evaluated the correlation between mtDNA alterations (mtDNA content, point mutations, large-scale deletions, and methylation status) and cisplatin sensitivity using two OSCC cell lines, namely SAS and H103, and stem cell-like tumour spheres derived from SAS. By microarray analysis, we found that the tumour spheres profited from aberrant lipid and glucose metabolism and became resistant to cisplatin. By qPCR analysis, we found that the cells with less mtDNA were less responsive to cisplatin (H103 and the tumour spheres). Based on the findings, we theorised that the metabolic changes in the tumour spheres probably resulted in mtDNA depletion, as the cells suppressed mitochondrial respiration and switched to an alternative mode of energy production, i.e. glycolysis. Then, to ascertain the origin of the variation in mtDNA content, we used MinION, a nanopore sequencer, to sequence the mitochondrial genomes of H103, SAS, and the tumour spheres. We found that the lower cisplatin sensitivity of H103 could have been caused by a constellation of genetic and epigenetic changes in its mitochondrial genome. Future work may look into how changes in mtDNA translate into an impact on cell function and therefore cisplatin response. Cis-diamminedichloroplatinum (II), or cisplatin, is one of the most commonly used chemotherapy agents in the treatment of various solid tumours such as ovarian, colorectal, prostate, lung, and head and neck tumours 1-5. To date, the intrinsic or acquired resistance of cancer cells to cisplatin remains a challenge in the chemotherapy of several cancers including oral squamous cell carcinoma (OSCC) 3,6. OSCC, which affects the epithelial layer of the oral cavity, is a common malignant tumour of the head and neck with low survival rates and high risks of recurrence 7. The well-characterized mode of action of cisplatin is via causing the formation of DNA adducts upon its binding to the nucleophilic N7 sites of purines, which further leads to DNA damage responses and apoptosis 2,6,8. Cisplatin resistance in general involves reduced DNA damage due to an increase in DNA adduct repair, reduced drug uptake, or increased drug inactivation 1,3,4,6. Activation of these mechanisms depends on multiple factors including genetic changes, epigenetic alterations at both molecular and cellular levels, and heterogeneity among cancer cells 4,9,10. The recently proposed cancer stem cells (CSCs) model highlighted tumour heterogeneity as an important basis of treatment resistance and relapse in cancer. According to the model, CSCs comprise a tumourigenic subpopulation where they exhibit stem cell-like features ...

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Section: Methodsmentioning

confidence: 99%

Mitochondrial DNA alterations may influence the cisplatin responsiveness of oral squamous cell carcinoma

Aminuddin

Leong

et al. 2020

Sci Rep

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“…We also tested a mtDNA deletion detection tool (MitoDel) in conjunction with two RNA-Seq alignment methods (MapSplice and STAR), as well as with the suggested DNA aligner (BWA), and compared these results to the sensitivities described above for our Splice-Break pipeline (31,34,36–38). Our assessment of the ‘common deletion’ demonstrated a robust effect of alignment method, with the poorest detection sensitivity (3–4%) resulting from MitoDel after BWA alignment (i.e.…”

Section: Resultsmentioning

confidence: 99%

“…Details about our rationale to use an RNA-Seq tool can be found in the online Supplemental Methods. We compared our process to an alternative mtDNA deletion detection method (MitoDel (31)) in addition to several alignment algorithms- MapSplice (34), TopHat (36), STAR (37) and BWA (38). The details of these comparisons can be found in the Results and online Supplemental Methods.…”

Section: Methodsmentioning

confidence: 99%

“…a variety of unknown breakpoints) would be hypothesized to occur in a tissue and thus would require a laborious workflow that includes plasmid cloning and selection (and/or single-cell isolation) in order to isolate and enrich each molecule sufficiently for Sanger sequencing (23,26). There have also been a few methods reported that use next-generation sequencing (NGS) data to identify mtDNA deletions; however, we believe these methods either lack vigor in regard to deletion quantification and breakpoint identification (29,30), or have been insufficiently tested with real mtDNA breakpoints and their associated repeat sequences (31). We have compared our bioinformatics pipeline to the MitoDel tool (31) using both DNA and RNA alignment algorithms in order to assess the effects of read mapping strategy and downstream filtering approaches on mtDNA deletion detection.…”

Section: Introductionmentioning

confidence: 99%

“…There have also been a few methods reported that use next-generation sequencing (NGS) data to identify mtDNA deletions; however, we believe these methods either lack vigor in regard to deletion quantification and breakpoint identification (29,30), or have been insufficiently tested with real mtDNA breakpoints and their associated repeat sequences (31). We have compared our bioinformatics pipeline to the MitoDel tool (31) using both DNA and RNA alignment algorithms in order to assess the effects of read mapping strategy and downstream filtering approaches on mtDNA deletion detection. Additional computational methods we have identified that may warrant future comparisons with our pipeline include a customized Perl script provided by Zambelli et al.…”

Section: Introductionmentioning

confidence: 99%

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Splice-Break: exploiting an RNA-seq splice junction algorithm to discover mitochondrial DNA deletion breakpoints and analyses of psychiatric disorders

Hjelm

Rollins

Morgan

et al. 2019

Nucleic Acids Research

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Deletions in the 16.6 kb mitochondrial genome have been implicated in numerous disorders that often display muscular and/or neurological symptoms due to the high-energy demands of these tissues. We describe a catalogue of 4489 putative mitochondrial DNA (mtDNA) deletions, including their frequency and relative read rate, using a combinatorial approach of mitochondria-targeted PCR, next-generation sequencing, bioinformatics, post-hoc filtering, annotation, and validation steps. Our bioinformatics pipeline uses MapSplice, an RNA-seq splice junction detection algorithm, to detect and quantify mtDNA deletion breakpoints rather than mRNA splices. Analyses of 93 samples from postmortem brain and blood found (i) the 4977 bp ‘common deletion’ was neither the most frequent deletion nor the most abundant; (ii) brain contained significantly more deletions than blood; (iii) many high frequency deletions were previously reported in MitoBreak, suggesting they are present at low levels in metabolically active tissues and are not exclusive to individuals with diagnosed mitochondrial pathologies; (iv) many individual deletions (and cumulative metrics) had significant and positive correlations with age and (v) the highest deletion burdens were observed in major depressive disorder brain, at levels greater than Kearns–Sayre Syndrome muscle. Collectively, these data suggest the Splice-Break pipeline can detect and quantify mtDNA deletions at a high level of resolution.

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Genomic Applications in Inherited Genetic Disorders

Krock¹,

Mao²,

Tvrdik³

et al. 2018

Genomic Applications in Pathology

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Detection and quantification of mitochondrial DNA deletions from next-generation sequence data

Cited by 31 publications

References 27 publications

Mitochondrial DNA alterations may influence the cisplatin responsiveness of oral squamous cell carcinoma

Mitochondrial DNA alterations may influence the cisplatin responsiveness of oral squamous cell carcinoma

Splice-Break: exploiting an RNA-seq splice junction algorithm to discover mitochondrial DNA deletion breakpoints and analyses of psychiatric disorders

Genomic Applications in Inherited Genetic Disorders

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