{
  "abstract": "GC-rich sequences are abundant in human herpesviruses genomes. GC-rich regions can form three-stranded RNA:DNA hybrid structures called R-loops. Though these hybrid structures serve important biological roles at telomeres or during cellular DNA synthesis, unscheduled or prolonged R-loop formation causes DNA damage and genome instability. For this reason, several mechanisms exist to resolve R-loops including endoribonucleases RNaseH1 (constitutively expressed) and RNaseH2A (cell cycle-regulated) which degrade the RNA portion of the R-loop. The Kaposi's sarcoma-associated herpesvirus (KSHV) origins of lytic replication (OriLyts) contain multiple cis-acting elements that are required for viral DNA replication including the production of GC-rich and repetitive transcripts, T1.4 (OriLyt-L) and kaposin (OriLyt-R). We previously showed that R-loops form at both OriLyts and that deleting kaposin repeats or decreasing their GC-rich content prevented R-loop formation at OriLyt-R, reduced genome amplification after primary infection and caused defects in latency establishment. To define the contribution that R-loops play in KSHV replication, we overexpressed RNaseH1, reasoning that excess RNaseH1 would resolve both OriLyt R-loops. However, RNaseH1 protein levels decreased following KSHV reactivation in both iSLK and BCBL-1 cell lines. Using co-transfection, we discovered that the KSHV viral replication and transcription activator protein, RTA, mediated RNaseH1 protein decreases in a E3 ligase domain-dependent manner without impacting levels of its cognate RNA transcript. We attempted to construct an RTA-resistant yet functional version of RNaseH1 by site-directed mutagenesis of lysine residues individually or in combination, yet these constructs remain susceptible to RTA-mediated protein decreases. An amino terminally tagged RNaseH1 displayed reduced susceptibility to RTA, suggesting that RTA may target the N-terminus of RNaseH1 for ubiquitination. However, overexpression of the cell-cycle regulated endonuclease, RNaseH2, exhibited RTA resistance, suggesting RNaseH2 may be a tool that will effectively resolve R-loops during KSHV infection. KSHV is not the only herpesvirus to encode a protein that reduces RNaseH1 levels, as co-expression of RTA homologs from the related gamma-herpesviruses EBV and MHV-68 likewise decreased steady-state levels of RNaseH1 protein. We propose that RTA-mediated RNaseH1 degradation is conserved strategy to ensure R-loop persistence during gamma-herpesvirus infection, underscoring the importance of these structures.",
  "abstract_shown": true,
  "affiliations": [
    "University of Calgary"
  ],
  "article_type": "New results",
  "arxiv": null,
  "author_total": 3,
  "authors": [
    {
      "name": "M. Kleer"
    },
    {
      "name": "J. Fox"
    },
    {
      "affs": [
        0
      ],
      "name": "J. Corcoran"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/ccywnqzn1ep9t.bib",
  "cited_by": 0,
  "doi": "10.64898/2026.08.31.748444",
  "fulltext_url": "https://www.biorxiv.org/content/10.64898/2026.08.31.748444v1.full",
  "id": "ccywnqzn1ep9t",
  "issns": [],
  "landing_url": "https://www.biorxiv.org/content/10.64898/2026.08.31.748444v1",
  "language": null,
  "licence": "CC-BY-NC-ND",
  "licence_label": "CC BY-NC-ND",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by-nc-nd/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "biorxiv",
      "microbiology"
    ],
    [
      "openalex",
      "2730|Viral-associated cancers and disorders"
    ]
  ],
  "openalex": "W7205005898",
  "pdf_url": "https://www.biorxiv.org/content/10.64898/2026.08.31.748444v1.full.pdf",
  "pmcid": null,
  "pmid": null,
  "published": "2026-08-31",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/ccywnqzn1ep9t.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:18:27Z",
      "native_id": "10.64898/2026.08.31.748444",
      "source": "biorxiv",
      "url": "https://www.biorxiv.org/content/10.64898/2026.08.31.748444v1"
    }
  ],
  "subjects": [
    {
      "field": "biology",
      "origin": "biorxiv:microbiology",
      "subfield": "microbiology"
    }
  ],
  "title": "Kaposi's sarcoma-associated herpesvirus forms and maintains R-loops at origins of lytic replication",
  "title_text": "Kaposi's sarcoma-associated herpesvirus forms and maintains R-loops at origins of lytic replication",
  "type": "preprint",
  "updated_at": "2026-10-30T12:18:27Z",
  "url": "https://research.draconitic.com/paper/ccywnqzn1ep9t",
  "venue": "bioRxiv",
  "versions": []
}