scholarly journals Single-cell whole-genome sequencing reveals mutational landscapes of DNA mismatch repair deficiency in mouse primary fibroblasts

2019 ◽  
Author(s):  
Lei Zhang ◽  
Xiao Dong ◽  
Xiaoxiao Hao ◽  
Moonsook Lee ◽  
Zhongxuan Chi ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Single Cell ◽  
Mismatch Repair ◽  
Genome Sequencing ◽  
Somatic Mutations ◽  
Fold Increase ◽  
Whole Genome ◽  
Wild Type ◽  
Dna Mismatch ◽  
Mmr Deficiency

AbstractDNA Mismatch repair (MMR) deficiency is a major cause of hereditary non-polyposis colorectal cancer, and is also associated with increased risk of several other cancers. This is generally ascribed to the role of MMR in avoiding mutations by correcting DNA replication errors. In MMR knockout mice very high frequencies of somatic mutations, up until 100-fold of background, have been reported. However, these results have been obtained using bacterial reporter transgenes, which are not representative for the genome overall, and mutational patterns of MMR deficiency remain largely unknown. To fill this knowledge gap, we performed single-cell whole-genome sequencing of lung fibroblasts of Msh2−/− and wild-type mice. We observed a 4-fold increase of somatic single nucleotide variants (SNVs) in the fibroblasts of Msh2−/− mice compared to those of wild-type mice. The SNV signature of Msh2 deficiency was found to be driven by C>T and T>C transitions. By comparing it to human cancer signatures, we not only confirmed the inferred MMR-deficiency-related etiology of several cancer signatures but also suggested that MMR deficiency is likely the cause of a cancer signature with its etiology previously unknown. We also observed a 7-fold increase of somatic small insertions and deletions (INDELs) in the Msh2−/− mice. An elevated INDEL frequency has also been found in human MMR-related cancers. INDELs and SNVs distributed differently across genomic features in the Msh2−/− and control cells, with evidence of selection pressure and repair preference. These results provide insights into the landscape of somatic mutations in normal somatic cells caused by MMR deficiency.SignificanceOur results show that MMR deficiency in the mouse is associated with a much lower elevation of somatic mutation rates than previously reported and provides the first MMR whole-genome mutational landscapes in normal somatic cells in vivo.

scholarly journals Determinants of Base-Pair Substitution Patterns Revealed by Whole-Genome Sequencing of DNA Mismatch Repair DefectiveEscherichia coli

Genetics ◽  
2018 ◽  
Vol 209 (4) ◽  
pp. 1029-1042 ◽  
Author(s):  
Patricia L. Foster ◽  
Brittany A. Niccum ◽  
Ellen Popodi ◽  
Jesse P. Townes ◽  
Heewook Lee ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Mismatch Repair ◽  
Genome Sequencing ◽  
Base Pair ◽  
Dna Mismatch Repair ◽  
Whole Genome ◽  
Dna Mismatch ◽  
Base Pair Substitution

scholarly journals Single-cell whole-genome sequencing reveals the functional landscape of somatic mutations in B lymphocytes across the human lifespan

2019 ◽  
Author(s):  
Lei Zhang ◽  
Xiao Dong ◽  
Moonsook Lee ◽  
Alexander Y. Maslov ◽  
Tao Wang ◽  
...  
Keyword(s):  
B Cell ◽  
Whole Genome Sequencing ◽  
Single Cell ◽  
B Lymphocytes ◽  
Genome Sequencing ◽  
Somatic Mutations ◽  
Somatic Hypermutation ◽  
Whole Genome ◽  
Healthy Individuals ◽  
Human Lifespan

Introductory paragraphThe accumulation of mutations in somatic cells have been implicated as a cause of ageing since the 1950s1,2. Yet, attempts to establish a causal relationship between somatic mutations and ageing have been constrained by the lack of methods to directly identify mutational events in primary human tissues. Here we provide detailed, genome-wide mutation frequencies and spectra of human B lymphocytes from healthy individuals across the entire human lifespan, from newborns to centenarians, using a recently developed, highly accurate single-cell whole-genome sequencing method3. We found that the number of somatic mutations increases from <500 per cell in newborns to >3,000 per cell in centenarians. We discovered mutational hotspot regions, some of which, as expected, located at immunoglobulin genes associated with somatic hypermutation. B cell-specific mutation signatures were observed associated with development, ageing or somatic hypermutation (SHM). The SHM signature strongly correlated with the signature found in human chronic lymphocytic leukemia and malignant B-cell lymphomas4, indicating that even in B cells of healthy individuals the potential cancer-causing events are already present. We also identified multiple mutations in sequence features relevant to cellular function, i.e., transcribed genes and gene regulatory regions. Such mutations increased significantly during ageing, but only at approximately half the rate of the genome average, indicating selection against mutations that impact B cell function. This first full characterization of the landscape of somatic mutations in human B lymphocytes indicates that spontaneous somatic mutations accumulating with age can be deleterious and may contribute to both the increased risk for leukemia and the functional decline of B lymphocytes in the elderly.

scholarly journals Determinants of base-pair substitution patterns revealed by whole-genome sequencing of DNA mismatch repair defectiveEscherichia coli

2018 ◽  
Author(s):  
Patricia L. Foster ◽  
Brittany A. Niccum ◽  
Ellen Popodi ◽  
Jesse P. Townes ◽  
Heewook Lee ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Mismatch Repair ◽  
Genome Sequencing ◽  
Base Pair ◽  
Whole Genome ◽  
Dna Mismatch ◽  
In The Wild ◽  
Leading Strand ◽  
Base Pair Substitution ◽  
Lagging Strand

ABSTRACTMismatch repair (MMR) is a major contributor to replication fidelity, but its impact varies with sequence context and the nature of the mismatch. Mutation accumulation experiments followed by whole-genome sequencing of MMR-defectiveE. colistrains yielded ≈30,000 base-pair substitutions, revealing mutational patterns across the entire chromosome. The base-pair substitution spectrum was dominated by A:T > G:C transitions, which occurred predominantly at the center base of 5′NAC3′+5′GTN3′ triplets. Surprisingly, growth on minimal medium or at low temperature attenuated these mutations. Mononucleotide runs were also hotspots for base-pair substitutions, and the rate at which these occurred increased with run length. Comparison with ≈2000 base-pair substitutions accumulated in MMR-proficient strains revealed that both kinds of hotspots appeared in the wild-type spectrum and so are likely to be sites of frequent replication errors. In MMR-defective strains transitions were strand biased, occurring twice as often when A and C rather than T and G were on the lagging-strand template. Loss of nucleotide diphosphate kinase increases the cellular concentration of dCTP, which resulted in increased rates of mutations due to misinsertion of C opposite A and T. In anmmr ndkdouble mutant strain, these mutations were more frequent when the template A and T were on the leading strand, suggesting that lagging-strand synthesis was more error-prone or less well corrected by proofreading than was leading strand synthesis.

scholarly journals Single-cell whole-genome sequencing reveals the functional landscape of somatic mutations in B lymphocytes across the human lifespan

2019 ◽  
Vol 116 (18) ◽  
pp. 9014-9019 ◽  
Author(s):  
Lei Zhang ◽  
Xiao Dong ◽  
Moonsook Lee ◽  
Alexander Y. Maslov ◽  
Tao Wang ◽  
...  
Keyword(s):  
B Cell ◽  
Whole Genome Sequencing ◽  
Single Cell ◽  
B Lymphocytes ◽  
Genome Sequencing ◽  
Somatic Mutations ◽  
Whole Genome ◽  
Healthy Individuals ◽  
Human Lifespan ◽  
Increased Risk

Accumulation of mutations in somatic cells has been implicated as a cause of aging since the 1950s. However, attempts to establish a causal relationship between somatic mutations and aging have been constrained by the lack of methods to directly identify mutational events in primary human tissues. Here we provide genome-wide mutation frequencies and spectra of human B lymphocytes from healthy individuals across the entire human lifespan using a highly accurate single-cell whole-genome sequencing method. We found that the number of somatic mutations increases from <500 per cell in newborns to >3,000 per cell in centenarians. We discovered mutational hotspot regions, some of which, as expected, were located at Ig genes associated with somatic hypermutation (SHM). B cell–specific mutation signatures associated with development, aging, or SHM were found. The SHM signature strongly correlated with the signature found in human B cell tumors, indicating that potential cancer-causing events are already present even in B cells of healthy individuals. We also identified multiple mutations in sequence features relevant to cellular function (i.e., transcribed genes and gene regulatory regions). Such mutations increased significantly during aging, but only at approximately one-half the rate of the genome average, indicating selection against mutations that impact B cell function. This full characterization of the landscape of somatic mutations in human B lymphocytes indicates that spontaneous somatic mutations accumulating with age can be deleterious and may contribute to both the increased risk for leukemia and the functional decline of B lymphocytes in the elderly.

scholarly journals 5-fluorocytosine resistance is associated with hypermutation and alterations in capsule biosynthesis in Cryptococcus

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
R. Blake Billmyre ◽  
Shelly Applen Clancey ◽  
Lucy X. Li ◽  
Tamara L. Doering ◽  
Joseph Heitman
Keyword(s):  
Whole Genome Sequencing ◽  
Mismatch Repair ◽  
Genome Sequencing ◽  
Resistance Genes ◽  
Fungal Pathogen ◽  
Glucuronic Acid ◽  
Nucleotide Metabolism ◽  
Whole Genome ◽  
Dna Mismatch

AbstractPatients infected with the fungal pathogen Cryptococcus are most effectively treated with a combination of 5-fluorocytosine (5FC) and amphotericin B. 5FC acts as a prodrug, which is converted into toxic 5-fluorouracil (5FU) upon uptake into fungal cells. However, the pathogen frequently develops resistance through unclear mechanisms. Here we show that resistance to 5FC in Cryptococcus deuterogattii is acquired more frequently in isolates with defects in DNA mismatch repair that confer an elevated mutation rate. We use whole genome sequencing of 16 independent isolates to identify mutations associated with 5FC resistance in vitro. We find mutations in known resistance genes (FUR1 and FCY2) and in a gene UXS1, previously shown to encode an enzyme that converts UDP-glucuronic acid to UDP-xylose for capsule biosynthesis, but not known to play a role in 5FC metabolism. Mutations in UXS1 lead to accumulation of UDP-glucuronic acid and alterations in nucleotide metabolism, which appear to suppress toxicity of both 5FC and its toxic derivative 5FU.

scholarly journals Whole-Genome Sequencing Reveals Breast Cancers with Mismatch Repair Deficiency

2017 ◽  
Vol 77 (18) ◽  
pp. 4755-4762 ◽  
Author(s):  
Helen Davies ◽  
Sandro Morganella ◽  
Colin A. Purdie ◽  
Se Jin Jang ◽  
Elin Borgen ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Mismatch Repair ◽  
Genome Sequencing ◽  
Whole Genome ◽  
Breast Cancers ◽  
Mismatch Repair Deficiency ◽  
Repair Deficiency

scholarly journals MBRS-59. SINGLE-CELL WHOLE-GENOME SEQUENCING DISSECTS INTRA-TUMOURAL GENOMIC HETEROGENEITY AND CLONAL EVOLUTION IN CHILDHOOD MEDULLOBLASTOMA

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii408-iii408
Author(s):  
Marina Danilenko ◽  
Masood Zaka ◽  
Claire Keeling ◽  
Stephen Crosier ◽  
Rafiqul Hussain ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Single Cell ◽  
Genome Sequencing ◽  
Copy Number ◽  
Single Cell Analysis ◽  
Mutational Analysis ◽  
Single Cells ◽  
Clonal Evolution ◽  
Whole Genome ◽  
Clinically Significant

Abstract Medulloblastomas harbor clinically-significant intra-tumoral heterogeneity for key biomarkers (e.g. MYC/MYCN, β-catenin). Recent studies have characterized transcriptional heterogeneity at the single-cell level, however the underlying genomic copy number and mutational architecture remains to be resolved. We therefore sought to establish the intra-tumoural genomic heterogeneity of medulloblastoma at single-cell resolution. Copy number patterns were dissected by whole-genome sequencing in 1024 single cells isolated from multiple distinct tumour regions within 16 snap-frozen medulloblastomas, representing the major molecular subgroups (WNT, SHH, Group3, Group4) and genotypes (i.e. MYC amplification, TP53 mutation). Common copy number driver and subclonal events were identified, providing clear evidence of copy number evolution in medulloblastoma development. Moreover, subclonal whole-arm and focal copy number alterations covering important genomic loci (e.g. on chr10 of SHH patients) were detected in single tumour cells, yet undetectable at the bulk-tumor level. Spatial copy number heterogeneity was also common, with differences between clonal and subclonal events detected in distinct regions of individual tumours. Mutational analysis of the cells allowed dissection of spatial and clonal heterogeneity patterns for key medulloblastoma mutations (e.g. CTNNB1, TP53, SMARCA4, PTCH1) within our cohort. Integrated copy number and mutational analysis is underway to establish their inter-relationships and relative contributions to clonal evolution during tumourigenesis. In summary, single-cell analysis has enabled the resolution of common mutational and copy number drivers, alongside sub-clonal events and distinct patterns of clonal and spatial evolution, in medulloblastoma development. We anticipate these findings will provide a critical foundation for future improved biomarker selection, and the development of targeted therapies.

Sign in / Sign up

Export Citation Format

Share Document