medRxiv (Cold Spring Harbor Laboratory), Aug 4, 2020
Background: Vast transcriptomics and epigenomics changes are characteristic of human cancers incl... more Background: Vast transcriptomics and epigenomics changes are characteristic of human cancers including leukemia. At remission, we assume that these changes normalise so that omics-profiles resemble those of healthy individuals. However, an in-depth transcriptomic and epigenomic analysis of cancer remission has not been undertaken. A striking exemplar of targeted remission induction occurs in chronic myeloid leukemia (CML) following tyrosine kinase inhibitor (TKI) therapy. Methods: Using RNA sequencing and whole-genome bisulfite sequencing, we profiled samples from chronic-phase CML patients at diagnosis and remission, and compared these to healthy donors. Results: Remarkably, our analyses revealed that abnormal splicing distinguishes remission samples from normal controls. This phenomenon is independent of the TKI drug used and in striking contrast to the normalisation of gene expression and DNA methylation patterns. Most remarkable are the high intron retention (IR) levels that even exceed those observed in the diagnosis samples. Increased IR affects cell cycle regulators at diagnosis and splicing regulators at remission. We show that aberrant splicing in CML is associated with reduced expression of specific splicing factors, histone modifications and reduced DNA methylation. Conclusions: Our results provide novel insights into the changing transcriptomic and epigenomic landscapes of CML patients during remission. The conceptually unanticipated observation of widespread aberrant alternative splicing after remission induction warrants further exploration. These results have broad implications for studying CML relapse and treating minimal residual disease.
Additional file 2. Figure 1: Box plot showing the difference in coverage across CpG islands, CpG ... more Additional file 2. Figure 1: Box plot showing the difference in coverage across CpG islands, CpG shores and other regions of the genome for each of the five library preparation methods compared. Figure 2 a, b Two representative examples of regions showing SNP in both the WGS and WGBS data of the same clinical sample. Figure 3 a Bar plot showing the percentage of SNPs from WGBS concordant in WGS-GS at ~ 26× coverage and the percentage of SNPs from spike-in WGS concordant in WGS-GS at 30× coverage. b A representative Venn diagram for one prostate cancer sample, 2ab showing the number of SNPs concordant at 26× coverage for WGBS and 30× coverage for spike-in WGS when compared with WGS-GS data. Figure 4 a, Plot showing the distribution of normalised frequency of number of SNPs called across regions of the genome with varying levels of GC content for WGBS and WGS-GS. b Plot showing the distribution of normalised frequency of number of SNPs called across regions of the genome with varying ...
Three-dimensional (3D) epigenome remodelling is an important mechanism of gene deregulation in ca... more Three-dimensional (3D) epigenome remodelling is an important mechanism of gene deregulation in cancer. However, its potential as a target to overcome therapy resistance remains largely unaddressed.Here we show that FDA-approved epigenetic therapy Decitabine (5-Aza-mC) suppresses tumour growth in preclinical metastatic ER+ breast tumour xenograft models. Decitabine-induced genome-wide DNA hypomethylation results in large-scale 3D epigenome deregulation, including de-compaction of higher order chromatin structure and loss of topologically associated domain boundary insulation. Significant DNA hypomethylation at ER-enhancer elements was associated with gain in ER binding, creation of ectopic 3D enhancer-promoter interactions and concordant activation of ER-mediated transcription pathways. Importantly long-term withdrawal of epigenetic therapy partially restores methylation at ER-enhancer elements, resulting in loss of ectopic 3D enhancer-promoter interactions and associated gene repres...
DNA methylation is required to maintain both DNA replication timing precision and 3D genome organ... more DNA methylation is required to maintain both DNA replication timing precision and 3D genome organization integrity Graphical abstract Highlights d DNA hypomethylation is associated with single-cell replication timing heterogeneity d Hypomethylation induces loss of allelic replication at cancerrelated gene loci d Hypomethylation alters higher-order genome architecture of PMD boundaries d Non-canonical H3K4me3-H3K9me3 domains form to protect silent late replication
Additional file 1:Â Table 1. Comparison of number of SNPs called in both WGBS and spike-in WGS da... more Additional file 1:Â Table 1. Comparison of number of SNPs called in both WGBS and spike-in WGS data at 13x and 30x coverage. Table 2. Comparison of number of SNPs concordant in spike-in WGS data with WGS-Gold Standard at 30x coverage. Table 3. Comparison of number of SNPs concordant in WGBS data with WGS-Gold Standard at 30x coverage. Table 4. Percentage of SNPs observed across different genomic contexts for WGS-GS and WGBS.
Vast transcriptomics and epigenomics changes are characteristic of human cancers, including leuka... more Vast transcriptomics and epigenomics changes are characteristic of human cancers, including leukaemia. At remission, we assume that these changes normalise so that omics-profiles resemble those of healthy individuals. However, an in-depth transcriptomic and epigenomic analysis of cancer remission has not been undertaken. A striking exemplar of targeted remission induction occurs in chronic myeloid leukaemia (CML) following tyrosine kinase inhibitor (TKI) therapy. Using RNA sequencing and whole-genome bisulfite sequencing, we profiled samples from chronic-phase CML patients at diagnosis and remission and compared these to healthy donors. Remarkably, our analyses revealed that abnormal splicing distinguishes remission samples from normal controls. This phenomenon is independent of the TKI drug used and in striking contrast to the normalisation of gene expression and DNA methylation patterns. Most remarkable are the high intron retention (IR) levels that even exceed those observed in t...
Monocytes and macrophages are essential components of the innate immune system. Herein, we report... more Monocytes and macrophages are essential components of the innate immune system. Herein, we report that intron retention (IR) plays an important role in the development and function of these cells. Using Illumina mRNA sequencing, Nanopore direct cDNA sequencing and proteomics analysis, we identify IR events that affect the expression of key genes/proteins involved in macrophage development and function. We demonstrate that decreased IR in nuclear-detained mRNA is coupled with increased expression of genes encoding regulators of macrophage transcription, phagocytosis and inflammatory signalling, including ID2, IRF7, ENG and LAT. We further show that this dynamic IR program persists during the polarisation of resting macrophages into activated macrophages. In the presence of proinflammatory stimuli, intron-retaining CXCL2 and NFKBIZ transcripts are rapidly spliced, enabling timely expression of these key inflammatory regulators by macrophages. Our study provides novel insights into the...
Endocrine therapy resistance frequently develops in estrogen receptor positive (ER+) breast cance... more Endocrine therapy resistance frequently develops in estrogen receptor positive (ER+) breast cancer, but the underlying molecular mechanisms are largely unknown. Here, we show that 3-dimensional (3D) chromatin interactions both within and between topologically associating domains (TADs) frequently change in ER+ endocrine-resistant breast cancer cells and that the differential interactions are enriched for resistance-associated genetic variants at CTCF-bound anchors. Ectopic chromatin interactions are preferentially enriched at active enhancers and promoters and ER binding sites, and are associated with altered expression of ER-regulated genes, consistent with dynamic remodelling of ER pathways accompanying the development of endocrine resistance. We observe that loss of 3D chromatin interactions often occurs coincidently with hypermethylation and loss of ER binding. Alterations in active A and inactive B chromosomal compartments are also associated with decreased ER binding and atypi...
HighlightsReplication timing alterations are conserved in cancers of different cell originsLong-r... more HighlightsReplication timing alterations are conserved in cancers of different cell originsLong-range epigenetic deregulation in cancer involves altered replication timingCancer late-replicating loci are hypomethylated and acquire facultative heterochromatinReplication timing status potentiates cis and trans chromosomal rearrangementsSummaryReplication timing is known to facilitate the establishment of epigenome, however, the intimate connection between DNA replication timing and changes to the genome and epigenome in cancer remain uncharted. Here, we perform Repli-Seq and integrated epigenome analysis and show that early-replicating loci are predisposed to hypermethylation and late-replicating loci to hypomethylation, enrichment of H3K27me3 and concomitant loss of H3K9me3. We find that altered replication timing domains correspond to long-range epigenetically deregulated regions in prostate cancer, and a subset of these domains are remarkably conserved across cancers from different...
Adipocytes support key metabolic and endocrine functions of adipose tissue. Lipid is stored in tw... more Adipocytes support key metabolic and endocrine functions of adipose tissue. Lipid is stored in two major classes of depots, namely visceral adipose (VA) and subcutaneous adipose (SA) depots. Increased visceral adiposity is associated with adverse health outcomes, whereas the impact of SA tissue is relatively metabolically benign. The precise molecular features associated with the functional differences between the adipose depots are still not well understood. Here, we characterised transcriptomes and methylomes of isolated adipocytes from matched SA and VA tissues of individuals with normal BMI to identify epigenetic differences and their contribution to cell type and depot-specific function. We found that DNA methylomes were notably distinct between different adipocyte depots and were associated with differential gene expression within pathways fundamental to adipocyte function. Most striking differential methylation was found at transcription factor and developmental genes. Our findings highlight the importance of developmental origins in the function of different fat depots. Adipocytes are the predominant cell type of adipose tissue and central to its key metabolic and endocrine functions, including storing or releasing triglycerides, and secreting adipokines that regulate metabolic, hormonal and inflammatory pathways 1,2. In mammals lipid is primarily stored in two major classes of depot, visceral adipose depots and subcutaneous adipose depots 2,3. While the different depots share many properties, there are important functional differences between visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT). These include differences in lipolysis, insulin sensitivity, adipokine secretion and inflammatory and immune function 4,5 .
Motivation: A synoptic view of the human genome benefits chiefly from the application of nucleic ... more Motivation: A synoptic view of the human genome benefits chiefly from the application of nucleic acid sequencing and microarray technologies. These platforms allow interrogation of patterns such as gene expression and DNA methylation at the vast majority of canonical loci, allowing granular insights and opportunities for validation of original findings. However, problems arise when validating against a "gold standard" measurement, since this immediately biases all subsequent measurements towards that particular technology or protocol. Since all genomic measurements are estimates, in the absence of a "gold standard" we instead empirically assess the measurement precision and sensitivity of a large suite of genomic technologies via a consensus modelling method called the row-linear model. This method is an application of the American Society for Testing and Materials Standard E691 for assessing interlaboratory precision and sources of variability across multiple testing sites. Both cross-platform and cross-locus comparisons can be made across all common loci, allowing identification of technology-and locus-specific tendencies. Results: We assess technologies including the Infinium MethylationEPIC BeadChip, whole genome bisulfite sequencing (WGBS), two different RNA-Seq protocols (PolyAþ and Ribo-Zero) and five different gene expression array platforms. Each technology thus is characterised herein, relative to the consensus. We showcase a number of applications of the row-linear model, including correlation with known interfering traits. We demonstrate a clear effect of cross-hybridisation on the sensitivity of Infinium methylation arrays. Additionally, we perform a true interlaboratory test on a set
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Papers by shalima nair