In This Section

RNA to the Rescue: A New DNA Repair Pathway for Double-Strand Breaks

Double-strand breaks (DSBs) represent one of the most deleterious forms of DNA damage, contributing to cancer development but also serving as key targets for therapies like radiation and certain chemotherapies. Traditional DNA repair pathways such as homologous recombination (HR), non-homologous end joining (NHEJ), and microhomology-mediated end joining (MMEJ) (1) are limited by their reliance on DNA templates and specific cell cycle phases. A study published in Nature Communications (2) by Manisha Jalan, DPhil, and colleagues reveal a novel RNA-templated DSB repair (RT-DSBR) pathway, mediated by the DNA polymerase ζ (Polζ) complex acting as a reverse transcriptase. Originally observed in in yeast (3), RT-DSBR in human cells challenges long-standing assumptions about reverse transcription and opens new possibilities for therapeutic innovation.

HR has long been the preferred method to deal with DSBs due to its error-free nature. Dr. Jalan hypothesized that in HR-deficient tumors, such as triple-negative breast cancer and ovarian cancer, cells must rely on alternative repair mechanisms to survive. With support from a 2020 AACR Swim Across America Cancer Research Fellowship, she investigated the role of RNA in guiding DNA repair and its functional consequences.

Manisha Jalan, DPhil

She and her team developed two complementary reporter assays, a fluorescence-based BFP-to-GFP conversion system and a sequencing-based AAVS1-seq assay, to directly detect RT-DSBR events. These assays demonstrated that RNA-containing oligonucleotides and mRNA transcripts can guide repair at CRISPR/Cas9-induced DSBs. Importantly, the study identified whole intron deletions (WIDs) in cancer genomes as a distinct mutational signature of RT-DSBR, suggesting that spliced RNA can serve as a template in vivo, occasionally leaving behind genomic scars. These findings expand the known repertoire of DNA repair pathways and raise new questions about the fidelity, regulation, and physiological relevance of RNA-guided repair.

Underscoring the broader implications of her work, Dr. Jalan noted that RT-DSBR may contribute to mutational and structural variations in tumors, particularly in transcriptionally active regions where RNA transcripts are abundant. The presence of WIDs in large cancer sequencing datasets suggests that this pathway plays a role in tumor evolution and genomic instability. Therapeutically, Polζ and its associated factors may emerge as novel targets in cancers that rely on RNA-templated repair for survival.

Unlike HR, which is restricted to dividing cells and requires a homologous DNA template, RT-DSBR may operate in non-dividing cells, such as neurons and muscle, where transcription is active but DNA templates are scarce. The discovery also opens new possibilities in gene editing and synthetic biology, where RNA-based donor templates could be engineered for precise genome modifications. At the same time, it introduces cautionary considerations for CRISPR applications, as off-target or unintended RNA-templated repair may occur, underscoring the need to better understand the regulation and fidelity of this pathway. Dr. Jalan notes that understanding the conditions under which RT-DSBR is activated, its error rates, and its regulatory mechanisms will be critical for assessing its impact on aging, neurodegeneration, and genome evolution.

Reflecting on the impact of the AACR grant on her research and career, Dr. Jalan shared: “This support allowed me to pursue innovative questions in the field of DNA repair, generating high-impact data that led to multiple publications and strengthened the foundation for future independent research. Beyond funding critical experiments, the AACR award provided invaluable visibility within the cancer research community, helping me establish new collaborations and refine my long-term research vision. This experience has greatly accelerated my transition toward independence, equipping me with the scientific rigor, leadership skills, and confidence needed to build a successful career in cancer research. I am forever grateful for the support.”

References

  1. Scully R, Panday A, Elango R, Willis NA. DNA double-strand break repair-pathway choice in somatic mammalian cells. Nat Rev Mol Cell Biol. 2019;20(11):698-714. doi:10.1038/s41580-019-0152-0
  2. Jalan M, Brambati A, Shah H, et al. RNA transcripts serve as a template for double-strand break repair in human cells. Nat Commun. 2025;16(1):4349. doi:10.1038/s41467-025-59510-x
  3. Meers C, Keskin H, Banyai G, et al. Genetic Characterization of Three Distinct Mechanisms Supporting RNA-Driven DNA Repair and Modification Reveals Major Role of DNA Polymerase ζ. Mol Cell. 2020;79(6):1037-1050.e5. doi:10.1016/j.molcel.2020.08.011