Inferring clonal somatic mutations directed by X chromosome inactivation status in single cells.
Ontology highlight
ABSTRACT:
SUBMITTER: Demirci I
PROVIDER: S-EPMC11312698 | biostudies-literature | 2024 Aug
REPOSITORIES: biostudies-literature
Publications
Inferring clonal somatic mutations directed by X chromosome inactivation status in single cells.
Demirci Ilke I Larsson Anton J M AJM Chen Xinsong X Hartman Johan J Sandberg Rickard R Frisén Jonas J
Genome biology 20240809 1
Analysis of clonal dynamics in human tissues is enabled by somatic genetic variation. Here, we show that analysis of mitochondrial mutations in single cells is dramatically improved in females when using X chromosome inactivation to select informative clonal mutations. Applying this strategy to human peripheral mononuclear blood cells reveals clonal structures within T cells that otherwise are blurred by non-informative mutations, including the separation of gamma-delta T cells, suggesting this ...[more]
Similar Datasets
Project description:BackgroundHigh-throughput sequencing allows the detection and quantification of frequencies of somatic single nucleotide variants (SNV) in heterogeneous tumor cell populations. In some cases, the evolutionary history and population frequency of the subclonal lineages of tumor cells present in the sample can be reconstructed from these SNV frequency measurements. But automated methods to do this reconstruction are not available and the conditions under which reconstruction is possible have not been described.ResultsWe describe the conditions under which the evolutionary history can be uniquely reconstructed from SNV frequencies from single or multiple samples from the tumor population and we introduce a new statistical model, PhyloSub, that infers the phylogeny and genotype of the major subclonal lineages represented in the population of cancer cells. It uses a Bayesian nonparametric prior over trees that groups SNVs into major subclonal lineages and automatically estimates the number of lineages and their ancestry. We sample from the joint posterior distribution over trees to identify evolutionary histories and cell population frequencies that have the highest probability of generating the observed SNV frequency data. When multiple phylogenies are consistent with a given set of SNV frequencies, PhyloSub represents the uncertainty in the tumor phylogeny using a "partial order plot". Experiments on a simulated dataset and two real datasets comprising tumor samples from acute myeloid leukemia and chronic lymphocytic leukemia patients demonstrate that PhyloSub can infer both linear (or chain) and branching lineages and its inferences are in good agreement with ground truth, where it is available.ConclusionsPhyloSub can be applied to frequencies of any "binary" somatic mutation, including SNVs as well as small insertions and deletions. The PhyloSub and partial order plot software is available from https://github.com/morrislab/phylosub/.
| S-EPMC3922638 | biostudies-literature
Project description:The most important prognostic factor for atherosclerotic cardiovascular disease is age, independent of all other recognized risk factors. Recently, exome sequence analyses showed that somatic mutations in blood cells, a process termed clonal hematopoiesis, are common and increase in prevalence with age, with at least 1 in 10 adults older than 70 years affected. Carriers of clonal hematopoiesis have been shown to be not only at heightened risk for hematologic malignancy but also at increased risk for atherosclerotic cardiovascular disease. Here, we review the prior literature of clonal selection and expansion of hematopoietic stem cells and the evidence supporting its causal association with atherosclerotic cardiovascular disease.
| S-EPMC6082163 | biostudies-literature
Project description:Accurate variant calling and genotyping represent major limiting factors for downstream applications of single-cell genomics. Here, we report Conbase for the identification of somatic mutations in single-cell DNA sequencing data. Conbase leverages phased read data from multiple samples in a dataset to achieve increased confidence in somatic variant calls and genotype predictions. Comparing the performance of Conbase to three other methods, we find that Conbase performs best in terms of false discovery rate and specificity and provides superior robustness on simulated data, in vitro expanded fibroblasts and clonal lymphocyte populations isolated directly from a healthy human donor.
| S-EPMC6444814 | biostudies-literature
Project description:During embryogenesis, the XIST RNA is expressed from and localizes to one X chromosome in females and induces chromosome-wide silencing. Although many changes to inactive X heterochromatin are known, the functional relationships between different modifications are not well understood, and studies of the initiation of X-inactivation have been largely confined to mouse. We now present a model system for human XIST RNA function in which induction of an XIST cDNA in somatic cells results in localized XIST RNA and transcriptional silencing. Chromatin immunoprecipitation and immunohistochemistry shows that this silencing need only be accompanied by a subset of heterochromatic marks and that these can differ between integration sites. Surprisingly, silencing is XIST-dependent, remaining reversible over extended periods. Deletion analysis demonstrates that the first exon of human XIST is sufficient for both transcript localization and the induction of silencing and that, unlike the situation in mice, the conserved repeat region is essential for both functions. In addition to providing mechanistic insights into chromosome regulation and formation of facultative heterochromatin, this work provides a tractable model system for the study of chromosome silencing and suggests key differences from mouse embryonic X-inactivation.
| S-EPMC1891207 | biostudies-literature
Project description:This unit describes a method of performing fluorescent in situ hybridization (FISH) of XIST and Cot-1 RNA in human pluripotent stem cells (hPSC) to characterize the epigenetic status of X-chromosome inactivation (XCI). hPSC laboratories commonly practice karyotypic analysis to monitor genetic stability; however, epigenetic stability is often overlooked. Several laboratories have recently shown that markers of XCI can be used as one effective screen to monitor the epigenetic status of hPSCs. Human embryonic stem cells (HESC) fall into three classes of XCI states: upregulating XIST upon differentiation, always expressing XIST in the undifferentiated and differentiated states, and never expressing XIST in the undifferentiated and differentiated states. Failure to express XIST represents an especially concerning state in hESC, as this state does not occur in healthy female cells but is often seen in malignancies. Herein, methods of carrying out XIST RNA and Cot-1 RNA FISH are described. FISH analysis of XIST RNA, unlike general expression analysis such as RT-PCR, allows for the classification of XCI on a single-cell level, enabling a quantitative determination of the degree of epigenetic change across the population. The complementary Cot-1 analysis measures the extent of repeat element expression throughout the nucleus and therefore enables determination, at a cytological level, of the extent to which the X chromosome is silent. Because the different steps of XCI are some of the first epigenetic changes to take place in differentiating hESC, analysis of the XCI state provides a first indication of an hESC culture's overall health.
| S-EPMC3741660 | biostudies-literature
Project description:Mammalian X chromosome inactivation (XCI) is an essential mechanism to compensate for dosage imbalances between male and female embryos. Although the molecular pathways are not fully understood, heterochromatinization of the Xi requires the coordinate recruitment of multiple epigenetic marks. Using fluorescence in situ hybridization analysis combined with immunocytochemistry, we demonstrate that the chromatin remodeling protein ATRX decorates the chromatids of a single, late replicating X chromosome in female somatic cells and co-localizes with the bona fide marker of the Xi, macroH2A1.2. Chromatin immunoprecipitation using somatic, embryonic stem (ES) cells and trophoblast stem (TS) cells as model for random and imprinted XCI, respectively, revealed that, in somatic and TS cells, ATRX exhibits a specific association with sequences located within the previously described H3K9me2-hotspot, a region 5' to the X inactive-specific transcript (Xist) locus. While no ATRX-Xi interaction was detectable in undifferentiated ES cells, an enrichment of ATRX was observed after 8 days of differentiation, indicating that ATRX associates with the Xi following the onset of random XCI, consistent with a potential role in maintenance of XCI. These results have important implications regarding a previously described escape from imprinted XCI in ATRX-deficient mice as well as cases of skewed XCI in patients with ATRX syndrome.
| S-EPMC2877807 | biostudies-literature
Project description:BACKGROUND:In patients with acquired aplastic anemia, destruction of hematopoietic cells by the immune system leads to pancytopenia. Patients have a response to immunosuppressive therapy, but myelodysplastic syndromes and acute myeloid leukemia develop in about 15% of the patients, usually many months to years after the diagnosis of aplastic anemia. METHODS:We performed next-generation sequencing and array-based karyotyping using 668 blood samples obtained from 439 patients with aplastic anemia. We analyzed serial samples obtained from 82 patients. RESULTS:Somatic mutations in myeloid cancer candidate genes were present in one third of the patients, in a limited number of genes and at low initial variant allele frequency. Clonal hematopoiesis was detected in 47% of the patients, most frequently as acquired mutations. The prevalence of the mutations increased with age, and mutations had an age-related signature. DNMT3A-mutated and ASXL1-mutated clones tended to increase in size over time; the size of BCOR- and BCORL1-mutated and PIGA-mutated clones decreased or remained stable. Mutations in PIGA and BCOR and BCORL1 correlated with a better response to immunosuppressive therapy and longer and a higher rate of overall and progression-free survival; mutations in a subgroup of genes that included DNMT3A and ASXL1 were associated with worse outcomes. However, clonal dynamics were highly variable and might not necessarily have predicted the response to therapy and long-term survival among individual patients. CONCLUSIONS:Clonal hematopoiesis was prevalent in aplastic anemia. Some mutations were related to clinical outcomes. A highly biased set of mutations is evidence of Darwinian selection in the failed bone marrow environment. The pattern of somatic clones in individual patients over time was variable and frequently unpredictable. (Funded by Grant-in-Aid for Scientific Research and others.).
| S-EPMC7478337 | biostudies-literature
Project description:Severe congenital neutropenia (SCN) and Shwachman-Diamond syndrome (SDS) are congenital neutropenia syndromes with a high rate of leukemic transformation. Hematopoietic stressors may contribute to leukemic transformation by increasing the mutation rate in hematopoietic stem/progenitor cells (HSPCs) and/or by promoting clonal hematopoiesis. We sequenced the exome of individual hematopoietic colonies derived from 13 patients with congenital neutropenia to measure total mutation burden and performed error-corrected sequencing on a panel of 46 genes on 80 patients with congenital neutropenia to assess for clonal hematopoiesis. An average of 3.6 ± 1.2 somatic mutations per exome was identified in HSPCs from patients with SCN compared with 3.9 ± 0.4 for healthy controls (P = NS). Clonal hematopoiesis due to mutations in TP53 was present in 48% (13/27) of patients with SDS but was not seen in healthy controls (0/17, P < .001) or patients with SCN (0/40, P < .001). Our SDS cohort was young (median age 6.3 years), and many of the patients had multiple TP53 mutations. Conversely, clonal hematopoiesis due to mutations of CSF3R was present in patients with SCN but was not detected in healthy controls or patients with SDS. These data show that hematopoietic stress, including granulocyte colony-stimulating factor, do not increase the mutation burden in HSPCs in congenital neutropenia. Rather, distinct hematopoietic stressors result in the selective expansion of HSPCs carrying specific gene mutations. In particular, in SDS there is enormous selective pressure to expand TP53-mutated HSPCs, suggesting that acquisition of TP53 mutations is an early, likely initiating event, in the transformation to myelodysplastic syndrome/acute myeloid leukemia in patients with SDS.
| S-EPMC5790127 | biostudies-literature
Project description:The Hi-C experiment can capture the genome-wide spatial proximities of the DNA, based on which it is possible to computationally reconstruct the three-dimensional (3D) structures of chromosomes. The transcripts of the long non-coding RNA (lncRNA) Xist spread throughout the entire X-chromosome and alter the 3D structure of the X-chromosome, which also inactivates one copy of the two X-chromosomes in a cell. The Hi-C experiments are expensive and time-consuming to conduct, but the Hi-C data of the active and inactive X-chromosomes are available. However, the Hi-C data of the X-chromosome during the process of X-chromosome inactivation (XCI) are not available. Therefore, the 3D structure of the X-chromosome during the process of X-chromosome inactivation (XCI) remains to be unknown. We have developed a new approach to reconstruct the 3D structure of the X-chromosome during XCI, in which the chain of DNA beads representing a chromosome is stored and simulated inside a 3D cubic lattice. A 2D Gaussian function is used to model the zero values in the 2D Hi-C contact matrices. By applying simulated annealing and Metropolis-Hastings simulations, we first generated the 3D structures of the X-chromosome before and after XCI. Then, we used Xist localization intensities on the X-chromosome (RAP data) to model the traveling speeds or acceleration between all bead pairs during the process of XCI. The 3D structures of the X-chromosome at 3 hours, 6 hours, and 24 hours after the start of the Xist expression, which initiates the XCI process, have been reconstructed. The source code and the reconstructed 3D structures of the X-chromosome can be downloaded from http://dna.cs.miami.edu/3D-XCI/.
| S-EPMC7772933 | biostudies-literature
Project description:TP53 protects cells from transformation by responding to stresses including aneuploidy and DNA double-strand breaks (DSBs). TP53 induces apoptosis of lymphocytes with persistent DSBs at antigen receptor loci and other genomic loci to prevent these lesions from generating oncogenic translocations. Despite this critical function of TP53, germline Tp53(-/-) mice succumb to immature T-cell (thymic) lymphomas that exhibit aneuploidy and lack clonal translocations. However, Tp53(-/-) mice occasionally develop B lineage lymphomas and Tp53 deletion in pro-B cells causes lymphomas with oncogenic immunoglobulin (Ig) locus translocations. In addition, human lymphoid cancers with somatic TP53 inactivation often harbor oncogenic IG or T-cell receptor (TCR) locus translocations. To determine whether somatic Tp53 inactivation unmasks translocations or alters the frequency of B lineage tumors in mice, we generated and analyzed mice with conditional Tp53 deletion initiating in hematopoietic stem cells (HSCs) or in lineage-committed thymocytes. Median tumor-free survival of each strain was similar to the lifespan of Tp53(-/-) mice. Mice with HSC deletion of Tp53 predominantly succumbed to thymic lymphomas with clonal translocations not involving Tcr loci; however, these mice occasionally developed mature B-cell lymphomas that harbored clonal Ig translocations. Deletion of Tp53 in thymocytes caused thymic lymphomas with aneuploidy and/or clonal translocations, including oncogenic Tcr locus translocations. Our data demonstrate that the developmental stage of Tp53 inactivation affects karyotypes of lymphoid malignancies in mice where somatic deletion of Tp53 initiating in thymocytes is sufficient to cause thymic lymphomas with oncogenic translocations.
| S-EPMC3885641 | biostudies-literature