Rapchak, K. | Search bioRxiv

bioRxiv2026

Brf1-Mediated RNA Polymerase III Activity Limits Murine Gammaherpesvirus Spread

RNA polymerase III (Pol III) activity is upregulated during herpesvirus infection, yet the functional consequences of this response remain poorly understood. To investigate the role of host Pol III transcription during murine gammaherpesvirus 68 (MHV68) infection, we depleted the Pol III transcription factor Brf1, an essential component of the TFIIIB complex required for transcription from Type I and Type II Pol III promoters. Brf1 depletion enhanced MHV68 replication during low multiplicity of infection (MOI) conditions, resulting in increased viral gene expression, viral protein accumulation, infectious virion production, and extracellular viral genome copies. These effects were confirmed to be Brf1-specific through rescue with an siRNA-resistant Brf1 construct. In contrast, Brf1 depletion produced only modest effects during high-MOI infection, suggesting that Brf1-dependent antiviral activity is most important during multistep viral spread. Transcriptomic analysis revealed accelerated induction of interferon-responsive genes early during infection in Brf1-deficient cells, followed by enhanced host transcript depletion at later stages, consistent with amplified host shutoff. Genetic disruption of the RIG-I/MAVS signaling pathway failed to abolish Brf1 antiviral activity, indicating that this phenotype is independent of MAVS-dependent interferon signaling. Finally, plaque assays demonstrated that Brf1-deficient cells supported larger plaques and increased plaque numbers, suggesting enhanced viral spread and/or entry. Together, our results reveal an unexpected role for Brf1-dependent Pol III activity in controlling gammaherpesvirus spread and suggest that virus-induced Pol III activation contributes to host antiviral defense.

microbiology↗

bioRxiv2022

Hrq1/RECQL4 regulation is critical for preventing aberrant recombination during DNA intrastrand crosslink repair and is upregulated in breast cancer.

Human RECQL4 is a member of the RecQ family of DNA helicases and functions during DNA replication and repair. RECQL4 mutations are associated with developmental defects and cancer. Although RECQL4 mutations lead to disease, RECQL4 overexpression is also observed in cancer, including breast and prostate. Thus, tight regulation of RECQL4 protein levels is crucial for genome stability. Because mammalian RECQL4 is essential, how cells regulate RECQL4 protein levels is largely unknown. Utilizing budding yeast, we investigated the RECQL4 homolog, HRQ1, during DNA crosslink repair. We find that Hrq1 functions in the error-free template switching pathway to mediate DNA intrastrand crosslink repair. Although Hrq1 repairs cisplatin-induced lesions, it is paradoxically degraded by the proteasome following cisplatin treatment. By identifying the targeted lysine residues, we show that preventing Hrq1 degradation results in increased recombination and mutagenesis. Like yeast, human RECQL4 is similarly degraded upon exposure to crosslinking agents. Furthermore, over-expression of RECQL4 results in increased RAD51 foci, which is dependent on its helicase activity. Using bioinformatic analysis, we observe that RECQL4 overexpression correlates with increased recombination and mutations. Overall, our study uncovers a role for Hrq1/RECQL4 in DNA intrastrand crosslink repair and provides further insight how misregulation of RECQL4 can promote genomic instability, a cancer hallmark.

genetics↗