Cellular DNA damage response pathways may be useful against some disease-causing viruses

Triggering a cell’s DNA damage response could be a promising avenue for developing new treatments for several rare but devastating viruses for which there are no antiviral treatments, including possibly the cancer-causing human papillomavirus (HPV), new research reveals.

Published online August 10 in Nucleic Acid ResearchThe paper focuses on the DNA damage response pathway and demonstrates how this pathway can reduce the function of a viral enzyme, a helicase, leading to suppression of viral replication.

“This research is important both for understanding how cells respond to DNA damage to prevent them from becoming cancerous in the first place, how targeting this pathway can be used in new cancer treatments, and because it now opens up possibilities for new approaches to treating some rare but devastating viral infections,” said Thomas Melendy, Ph.D., senior author of the paper and associate professor of microbiology and immunology in the Jacobs School of Medicine and Biomedical Sciences at the University at Buffalo.

How replication slows in response to DNA damage

The research focuses on a process called the DNA damage response, part of which evolved to stop or slow down DNA synthesis whenever damage to cellular DNA occurs.

“These pathways are important in preventing the exacerbation of DNA damage that can lead to either cell death or cancer,” says Rama Dey-Rao, Ph.D., research assistant professor of microbiology and immunology in the Jacobs School and co-senior author of the paper with Caleb Hominski, Ph.D., a former student in the lab.

When these pathways are activated, DNA replication is inhibited at sites in the genome called origins; at the same time, the progression of DNA replication forks also slows. Replication forks, so called because their structure resembles a fork, are where large groups of proteins coordinate genome replication through the unwinding and synthesis of DNA.

Melendy says that while much is known about how the DNA damage response causes cells to prevent DNA replication origins from “firing,” it is much more difficult to understand how replication fork progression slows in response to DNA damage.

“Researchers have been very interested in how this slowdown occurs because it is so dramatic,” Melendy says. “DNA damage response pathways cause the progression of replication forks to slow down by about tenfold. This tenfold slowdown means that the synthesis of the cell’s genome, which usually takes about 12 hours, would take almost five days, which dramatically increases the time that cells have to repair DNA damage.”

A viral connection

For years, Melendy and his colleagues have been studying two types of small DNA viruses: papillomaviruses, such as HPV, and polyomaviruses, which infect most people without causing serious illness but can lead to serious illness and some cancers in immunologically compromised individuals. A rare cancer caused by a polyomavirus killed musician Jimmy Buffett in 2023.

“We have previously shown that in response to DNA damage, HPV does not stop or slow down its DNA replication, whereas polyomaviruses stop or slow down their DNA replication,” Melendy says. “So by comparing and contrasting these two types of viruses, we can better understand how polyomavirus DNA replication is slowed down in response to DNA damage, which in turn gives us insight into how human cells slow down replication forks.”

In the current research, they demonstrate that a phosphorylation site (where a phosphate is added to a molecule) on the main DNA replication and transcription protein of the polyomavirus is highly conserved in polyomaviruses from many animal species.

“The conservation of this phosphorylation/modification across polyomaviruses that have evolved to infect many different mammalian species suggests that this was likely important,” Melendy says.

To study the effects of this phenomenon, the UB researchers made a mutation at the specific amino acid residue of the viral protein where this phosphorylation occurs to mimic the addition of a phosphate group present at that location.

By expressing this mutant viral protein in human cells using a polyomavirus DNA replication assay system, they found that replication of the virus genome was reduced by 10-fold. However, viral transcription was not affected, indicating that phosphorylation of this amino acid residue has a very specific effect on viral DNA replication, but did not affect other functions of this protein.

The role of DNA helicase

Comparing the wild-type and mutant proteins, they found that the only compromised function was the ability to act as a DNA helicase, unwinding DNA strands to facilitate the entry of DNA-synthesizing enzymes.

“This is the first demonstration that it might be possible to use phosphorylation as a ‘switch’ on a DNA helicase to reduce the speed of replication,” Melendy says.

Evidence suggests that similar phosphorylation may also occur in human DNA helicases.

“For many cancers, if we selectively inhibit the DNA damage checkpoints that they still retain, and simultaneously treat them with lower than normal amounts of DNA-damaging chemotherapies, then we may be able to selectively damage cancer cells while leaving non-cancer cells intact, thereby dramatically enhancing cancer cell killing while simultaneously reducing toxic side effects.”

This is an ongoing area of ​​study being conducted by UB researchers with their collaborators at Roswell Park Comprehensive Cancer Center.

Based on the current study, these DNA damage checkpoints could now be relevant for the treatment of viral infections of the small DNA viruses studied at UB.

“Because they rely almost exclusively on host cell enzymes to synthesize their viral genomes, these small DNA viruses have proven to be highly resistant to antiviral therapies,” Melendy says.

“We currently have no antiviral treatments for HPV or polyomaviruses. By triggering the DNA damage response in a patient, this could significantly slow down viral DNA replication, thereby suppressing the infection, providing us with a new avenue for potential antiviral treatments for these as yet incurable viral infections.”