March 03, 2025

How a crucial DNA repair protein works-and what it means for cancer treatment New structural blueprint is key for better targeting cancer cells, particularly those with BRCA1 and 2 mutations.

March 03, 2025

LA JOLLA, CA DNA repair proteins act like the body's editors, constantly finding and reversing damage to our genetic code. Researchers have long struggled to understand how cancer cells hijack one of these proteins called polymerase theta (Pol-theta) for their own survival. But scientists at Scripps Research have now captured the first detailed images of Pol-theta in action, revealing the molecular processes responsible for a range of cancers.

The findings, published in Nature Structural & Molecular Biology on February 28, 2025, illuminate how Pol-theta undergoes a major structural rearrangement when it binds to broken DNA strands. By unveiling Pol-theta's DNA-bound structure its active state the study provides a blueprint for designing more effective cancer drugs.

We now have a much clearer picture of how Pol-theta works, which will enable us to block its activity more precisely, says senior author Gabriel Lander, a professor at Scripps Research.

Technically, Pol-theta is an enzyme a type of protein that speeds up chemical reactions, including those related to cell repair. DNA damage is a constant problem for cells, occurring millions of times per day collectively throughout our bodies. Cells normally use highly accurate mechanisms to fix these breaks, but some cancers particularly those arising from BRCA1 or BRCA2 mutations, such as certain breast and ovarian cancers lack this function. Instead, they depend on a more error-prone method, controlled by Pol-theta.

Pol-theta is an important target, and many pharmaceutical companies see it as a promising way to treat cancers that have defective DNA repair pathways, adds first author Christopher Zerio, a former postdoctoral fellow in Lander's lab.

Although previous studies have mapped parts of Pol-theta's structure, the enzyme's interactions with DNA weren't well understood.

What's been missing is how Pol-theta actually engages DNA, which is essential for drug development, says Zerio.

Prior research has shown that Pol-theta exists in two forms: a tetramer (four copies of the enzyme) and a dimer (two copies). But why or how Pol-theta changed between these forms was unknown.

Before this study, Pol-theta's structure had only been captured in an inactive state, leaving a major knowledge gap regarding how the enzyme interacts with DNA. It was like trying to determine how a bee accesses nectar when all you've ever seen is a closed flower.

You know the interaction must happen, but without seeing it, the mechanism remains a mystery, explains Lander.

Using cryo-electron microscopy and biochemical experiments, the team made a surprising discovery while capturing Pol-theta in the act of repairing DNA: Whenever Pol-theta bound to broken strands, it consistently switched from the tetrameric to a never-before-seen dimeric configuration.

Once in its active state, Pol-theta repairs DNA using a two-step process: First, the enzyme searches for small matching sequences called microhomologies on broken strands. Once a matching sequence is found, Pol-theta holds the broken DNA strands together so that they can be stitched together without needing extra energy. Most enzymes require an energy boost to function, but Pol-theta relies on the natural attraction between matching DNA sequences, allowing them to snap into place on their own.

If we can block this process, we could make Pol-theta-dependent cancers much more sensitive to treatment, says Zerio.

Importantly, Pol-theta is produced at low levels in healthy cells, making it a promising target for cancer therapies. Unlike cancer cells, which depend on Pol-theta as a workaround for defective repair pathways, healthy ones rely on more accurate repair mechanisms that require energy ensuring more precise DNA repair. Because healthy cells don't need Pol-theta for survival, blocking the enzyme's activity likely won't cause widespread damage to healthy tissue.

Most cancer drugs target proteins that are also needed by healthy cells, notes Lander. Specifically targeting Pol-theta should only kill cancer cells, lowering the chance of side effects during therapy.

Drugs that inhibit Pol-theta are already in clinical trials, but they currently must be combined with other therapies to work effectively. While this study could inform more precise drug development, further research may reveal other roles the enzyme may play in cellular functions.

We also want to understand why Pol-theta exists in its tetrameric form and how it interacts with other DNA repair enzymes, says Lander. Such insights could lead to new ways of targeting BRCA-associated cancers.

In addition to Lander and Zerio, authors of the study, Human polymerase helicase positions DNA microhomologies for double-strand break repair, include Yonghong Bai, Brian A. Sosa-Alvarado and Timothy Guzi of MOMA Therapeutics.

This work was supported by funding from the National Institutes of Health (F32CA288144, GM14305 and S10OD032467).

Integrative Structural & Computational Biology Lander, Gabriel

LINK:	https://www.scripps.edu/news-and-events/press-room/2025/20250303-lande...
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