Biology subjects

Sokol, D.

Publications and source records attributed to Sokol, D..

3 recordsLinked to original sources

bioRxiv2024

High-coverage allele-resolved single-cell DNA methylation profiling by scDEEP-mC reveals cell lineage, X-inactivation state, and replication dynamics

DNA methylation is a relatively stable epigenetic mark with important roles in development and disease.1 Since cell-to-cell variation in epigenetic programming can reflect important differences in cell state and fate, it is clear that single-cell methods are essential to understanding this key epigenetic mark in heterogeneous tissues. Existing single-cell whole-genome bisulfite sequencing (scWGBS) methods2 have significant shortcomings, including very low CpG coverage3-7 or inefficient library generation requiring extremely deep sequencing.8 These methods offer limited insight into focal regulatory regions and generally preclude direct cell-to-cell comparisons. To address these shortcomings, we have developed an improved method, scDEEP-mC (single-cell Deep and Efficient Epigenomic Profiling of methyl-C). We show that high-coverage promoter methylation profiling by scDEEP-mC can identify multiple cell types, while allele-resolved methylation calls allow assessment of X-inactivation state in single cells and identification of transcription factor binding sites (TFBS) enriched for hemi-methylation. We also use scDEEP-mC to profile single-cell copy number alterations, identify actively replicating cells, and track DNA methylation dynamics during and after DNA replication. The high coverage of scDEEP-mC creates an exceptional opportunity to explore DNA methylation biology in individual cells.

genomics↗

bioRxiv2020

RNA-seq of Human T-Cells After Hematopoietic Stem Cell Transplantation Identifies Linc00402 as a Novel Regulator of T-Cell Alloimmunity

Mechanisms governing allogeneic T-cell responses after allogeneic hematopoietic stem cell (HSC) and solid organ transplantation are incompletely understood. Long non-coding RNAs (lncRNA) do not code for, but control gene expression with tissue specificity. However, their role in T-cell alloimmunity is unknown. We performed RNA-seq on donor T-cells from HSCT patients and found that increasing strength of allogeneic stimulation caused greater differential expression of lncRNAs. The differential expression was validated in an independent patient cohort, and also following ex vivo allogeneic stimulation of healthy human T-cells. Linc00402, a novel, conserved lncRNA, was identified as the most differentially expressed and was enriched 88 fold in human T-cells. Mechanistically, it was mainly located in the cytoplasm, and its expression was rapidly reduced following T-cell activation. Consistent with this, tacrolimus preserved the expression of Linc00402 following T-cell activation, and lower levels of Linc00402 were found in patients who subsequently went on to develop acute graft versus host disease (GVHD). The dysregulated expression of Linc00402 was also validated in murine T-cells, both in vitro and in vivo. Functional studies using multiple modalities to deplete Linc00402 in both mouse and human T-cells, demonstrated a critical role for Linc00402 in the T-cell proliferative response to an allogeneic stimulus but not a non-specific anti-CD3/CD28 stimulus. Thus, our studies identified Linc00402 as a novel, conserved regulator of allogeneic T-cell function. Because of its T-cell specific expression and its impact on allogeneic T-cell responses, targeting Linc00402 may improve outcomes after allogeneic HSC and solid organ transplantation. One sentence summaryLncRNAs are differentially expressed by allogeneic antigen-stimulated T-cells, and the novel lncRNA, Linc00402, is a specific regulator of mouse and human allogeneic T-cells.

immunology↗

bioRxiv2019

Altered synaptic ingestion by human microglia in Alzheimer’s disease

Synapse loss correlates with cognitive decline in Alzheimers disease (AD). Data from mouse models suggests microglia are important for synapse degeneration, but direct human evidence for any glial involvement in synapse removal in human AD remains to be established. Here we observe astrocytes and microglia from human brains contain greater amounts of synaptic protein in AD compared to non-disease controls, and that proximity to amyloid-{beta} plaques and the APOE4 risk gene exacerbate this effect. In culture, mouse and human astrocytes and primary mouse and human microglia phagocytose AD patient-derived synapses more than synapses from controls. Inhibiting MFG-E8 function rescued the elevated engulfment of AD synapses by astrocytes and microglia without affecting control synapse uptake. Thus, AD promotes increased synapse ingestion by human glial cells via an MFG-E8 opsonophagocytic mechanism with potential for targeted therapeutic manipulation. One-Sentence SummaryGlial cells ingest synapses in Alzheimers disease and antibody treatment reduces this ingestion in cultured human cells.

neuroscience↗