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Reconstructing EBV reactivation and DNA damage response kinetics in morphologic pseudotime.

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Isolation and Quantification of Epstein-Barr Virus from the P3HR1 Cell Line
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Reconstructing EBV reactivation and DNA damage response kinetics in morphologic pseudotime.

Dina G Tekle1, Jonathan Z Sexton2, Elliott D SoRelle1,3

  • 1Department of Biological Chemistry, University of Michigan, Ann Arbor, MI 48109.

Biorxiv : the Preprint Server for Biology
|November 24, 2025
PubMed
Summary
This summary is machine-generated.

Epstein-Barr virus (EBV) subverts host DNA damage responses (DDR) during lytic infection. This study reveals how EBV manipulates DDR proteins like 53BP1, impacting viral replication and host cell dynamics.

Keywords:
53BP1B cellDNA damage responseEpstein-Barr virusHigh-content screeningcell dynamicsgenotoxic stresshost-virus interactionsinfectionlymphomalytic reactivationmicroscopymorphologic profilingpseudotimesingle-cellyH2AX

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Area of Science:

  • Virology
  • Cell Biology
  • Immunology

Background:

  • Epstein-Barr virus (EBV) lytic infection is linked to cancers and autoimmune diseases.
  • EBV's lytic cycle relies on manipulating host DNA damage responses (DDR).
  • Understanding EBV-host interactions during lytic replication is crucial for therapeutic strategies.

Purpose of the Study:

  • To profile single-cell EBV reactivation and DDR dynamics using high-content screening (HCS).
  • To create a comprehensive atlas of cellular phenotypes during EBV lytic induction.
  • To investigate the spatial and temporal regulation of DDR factors during EBV replication.

Main Methods:

  • High-content screening (HCS) of EBV lytic induction across nine B cell lymphoma models.
  • Pseudotemporal profiling of single-cell EBV reactivation and DDR dynamics.
  • Single-cell segmentation, feature extraction, and clustering to analyze cellular responses.

Main Results:

  • Generated an atlas of >750,000 cells detailing lytic protein expression, DNA replication, and DDR factors.
  • Identified distinct treatment- and model-dependent cellular responses to EBV lytic induction.
  • Observed differential DDR profiles in lytic vs. latent cells and varied lytic protein localization.
  • Revealed depletion of γH2AX from viral replication compartments (VRCs) but widespread host chromatin localization.
  • Found 53BP1 present before viral replication but absent from VRCs and host chromatin during reactivation, indicating DDR dysregulation.

Conclusions:

  • EBV transiently utilizes host double-strand break (DSB) DDR mediators for initial genome replication.
  • EBV initiates but ultimately impairs host-targeted DDR during reactivation.
  • The study presents a powerful HCS technique for analyzing host-virus dynamics in single cells.
  • This method supports future high-throughput single-cell virology applications.