Topological Stress Induces Difficult-to-Repair DNA Lesions in Ribosomal DNA, Leading to the Formation of PML-Nucleolar Compartments

Core Concepts

Topological stress and inhibition of RNA polymerase I can induce double-strand breaks in the ribosomal DNA locus, leading to the formation of PML-nucleolar compartments that segregate damaged rDNA from active nucleoli.

Abstract

The study investigates the stimuli that trigger the formation of PML-nucleolar associations (PNAs), which are formed when the multifunctional protein PML associates with nucleolar caps after RNA polymerase I (RNAPI) inhibition. The researchers exposed cells to various genotoxic stresses and found that the most potent inducers of PNAs were those that introduced topological stress and inhibited RNAPI, such as the chemotherapeutic drug doxorubicin.
Doxorubicin was found to induce double-strand breaks (DSBs) specifically in the ribosomal DNA (rDNA) locus. The PNAs co-localized with the damaged rDNA, segregating it from active nucleoli. Cleaving the rDNA locus with the I-PpoI enzyme confirmed that rDNA damage is a genuine stimulus for PNA formation.
The study further showed that the formation of I-PpoI-induced PNAs depends on ATM/ATR-dependent nucleolar cap formation and homologous recombination (HR) repair pathways. The PNAs co-localized with rDNA DSBs that were positive for RPA32-pS33 (a marker of resected DNA) but deficient in RAD51 (a key HR protein), indicating that the DNA damage was unable to complete HR repair.
The findings suggest that PNAs form in response to persistent rDNA damage within the nucleolar cap, highlighting the interplay between PML/PNAs and rDNA alterations due to topological stress, RNAPI inhibition, and rDNA DSBs destined for HR. Cells with persistent PNAs undergo senescence, suggesting that PNAs help avoid rDNA instability, with implications for tumorigenesis and aging.

Stats

Doxorubicin, a chemotherapeutic drug, induced double-strand breaks (DSBs) in the ribosomal DNA (rDNA) locus.
Inhibition of ATM, ATR kinases, and RAD51 reduced I-PpoI-induced PML-nucleolar associations (PNAs).
I-PpoI-induced PNAs co-localized with rDNA DSBs positive for RPA32-pS33 but deficient in RAD51, indicating resected DNA unable to complete homologous recombination repair.

Quotes

"PNAs form in response to persistent rDNA damage within the nucleolar cap, highlighting the interplay between PML/PNAs and rDNA alterations due to topological stress, RNAPI inhibition, and rDNA DSBs destined for HR."
"Cells with persistent PNAs undergo senescence, suggesting PNAs help avoid rDNA instability, with implications for tumorigenesis and aging."

Key Insights Distilled From

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

by Hornofova,T.... at www.biorxiv.org 08-06-2023

https://www.biorxiv.org/content/10.1101/2023.08.05.552131v3

Deeper Inquiries

How do the PML-nucleolar compartments contribute to cellular senescence, and what are the potential implications for cancer and aging?

The formation of PML-nucleolar compartments (PNAs) plays a crucial role in cellular senescence by segregating damaged ribosomal DNA (rDNA) from active nucleoli. This segregation helps prevent rDNA instability, which can lead to genomic instability and cellular dysfunction. In the context of cancer, the presence of persistent PNAs can trigger senescence in cancer cells, acting as a protective mechanism against tumorigenesis. On the other hand, in aging cells, the accumulation of PNAs may contribute to the aging process by promoting senescence and limiting the proliferation of damaged cells. Therefore, understanding the role of PNAs in cellular senescence has significant implications for both cancer development and aging-related processes.

What other cellular processes or pathways might be affected by the segregation of damaged rDNA from active nucleoli?

The segregation of damaged rDNA from active nucleoli, facilitated by the formation of PML-nucleolar compartments (PNAs), can impact several cellular processes and pathways. One major consequence is the disruption of ribosomal RNA synthesis, which is essential for protein production and cell growth. The inhibition of RNA polymerase I (RNAPI) due to the presence of PNAs can lead to nucleolar stress and activation of stress response pathways such as the p53 pathway. Additionally, the segregation of damaged rDNA can affect nucleolar functions beyond ribosome biogenesis, including the regulation of cell cycle progression, DNA repair processes, and cellular signaling pathways. The altered nucleolar dynamics resulting from damaged rDNA segregation can have widespread effects on cellular homeostasis and function.

Could the insights from this study be leveraged to develop new therapeutic strategies targeting the PML-nucleolar response to topological stress and rDNA damage?

The findings from this study provide valuable insights into the mechanisms underlying the PML-nucleolar response to topological stress and rDNA damage, offering potential opportunities for the development of novel therapeutic strategies. Targeting the formation of PML-nucleolar compartments (PNAs) could be a promising approach to modulate cellular senescence, particularly in cancer cells where senescence induction can inhibit tumor growth. By understanding the signaling pathways and molecular interactions involved in PNAs formation, researchers may identify druggable targets that can disrupt or prevent the assembly of PNAs in response to topological stress and rDNA damage. Furthermore, targeting specific components of the PML-nucleolar response, such as ATM/ATR kinases or homologous recombination factors, could offer new avenues for therapeutic intervention in diseases associated with nucleolar stress and genomic instability. Leveraging the insights from this study could lead to the development of targeted therapies aimed at modulating the PML-nucleolar response for clinical applications in cancer treatment and age-related disorders.