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Topological stress triggers persistent DNA lesions in ribosomal DNA with ensuing formation of PML-nucleolar

Alexandra Urbancokova1, Terezie Hornofova1, Josef Novak1

  • 1Laboratory of Genome Integrity, Institute of Molecular Genetics of the Czech Academy of Sciences, Prague, Czech Republic.

Elife
|October 10, 2024
PubMed

Insights

PML-nucleolar associations (PNAs) form when cells experience DNA damage and RNA polymerase I inhibition. These structures help prevent genomic instability, potentially impacting aging and cancer.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Genetics

Background:

  • PML protein is essential for PML-nuclear bodies, which regulate cellular stress responses.
  • PML can associate with nucleolar caps, forming PML-nucleolar associations (PNAs), particularly after RNA polymerase I (RNAPI) inhibition.
  • The precise stimuli and mechanisms triggering PNAs remain incompletely understood.

Purpose of the Study:

  • To identify the specific cellular stresses that induce PNAs.
  • To elucidate the molecular mechanisms underlying PNA formation.
  • To investigate the functional consequences of PNAs in cellular response to DNA damage.

Main Methods:

  • Cells were exposed to various genotoxic stresses to identify PNA inducers.
  • Specific compounds like Doxorubicin and rDNA cleavage with I-PpoI were used to induce and study PNAs.
  • Inhibition of key kinases (ATM, ATR) and DNA repair proteins (RAD51) were employed to dissect the PNA pathway.
  • Immunofluorescence was used to assess co-localization of PNAs with damaged rDNA and DNA repair markers.

Main Results:

  • Topological stress and RNAPI inhibition were potent PNA inducers, with Doxorubicin inducing double-strand breaks (DSBs) at the rDNA locus.
  • PNAs co-localized with damaged rDNA, separating it from active nucleoli, and rDNA damage was confirmed as a direct stimulus.
  • PNA formation was dependent on ATM/ATR signaling and homologous recombination (HR) pathway components, with PNAs accumulating at resected but unrepaired rDNA DSBs.
  • Persistent PNAs correlated with cellular senescence, suggesting a role in preventing genomic instability.

Conclusions:

  • PNAs are a cellular response to persistent rDNA damage within nucleolar caps, triggered by topological stress, RNAPI inhibition, and rDNA DSBs requiring HR.
  • The formation of PNAs involves ATM/ATR-dependent pathways and the early stages of HR repair.
  • Persistent PNAs are linked to senescence, indicating a protective role against rDNA instability with implications for aging and tumorigenesis.
Keywords:
PMLaberrant DNA topologycancer biologycell biologycellular senescencegenome maintenancehumanpersistent rDNA damage

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