Researchers from Baylor College of Medicine (BCM) have developed a new tool that allows them to quantify how often dangerous double-stranded breaks in DNA occur. They used a bacterial virus protein known as Gam fused to a green fluorescent protein that has the ability to bind to these DNA areas. These dangerous double-stranded breaks are known to destabilize the genome, driving both evolution and cancer. Thus far, researchers have learned new things about how DNA breaks occur in bacterial, mouse and human cells.
Dr. Susan Rosenburg, professor of molecular and human genetics at BCM and corresponding author of the report, and colleagues, made use of this new tool to determine how spontaneous double-stranded breaks occur in bacterial and mammalian cells. Gam (viral protein) was used due to the fact that it binds to double-stranded DNA breaks. Gam doesn’t bind to other proteins or unbroken DNA. When Gam is fused with a green fluorescent tag (GamGFP) and introduced into cells, the double-stranded breaks light up green. Drs. Kyle M. Miller of the University of Texas at Austin, and Reuben S. Harris of the University of Minnesota were also corresponding authors.
The researchers estimated that the GamGFP detects between 71 percent and 82 percent double-stranded breaks in bacteria. Their results support the idea that many spontaneous double-stranded breaks occur during replication. Replication is when the DNA makes a copy of itself getting ready for cell division. Interestingly, they also discovered a type of DNA breakage that involves the innate immune system that is specific to primates including humans.
Dr. Michael Cox of the department of biochemistry at the University of Wisconsin-Madison in a corresponding viewpoint notes that there are limitations to the GamGFP technique, however the new technique is destined for creative applications in eukaryotic cell biology. As he points out, the list of potential experiments seems endless.
The GamGFP technique is part of developing innovative new tools, initiated and led by the Rosenberg lab. Their goal is to research proteins that cause DNA damage and might promote cancer. The lab is supported by a National Institutes of Health Director’s Pioneer Award.
Other contributors to this research include: Chandan Shee, Franklin Gu, Mohan C Joshi, David Magnan, Jennifer A Halliday, Ryan L. Frisch, Janet L Gibson, Ralf Bernd Nehring, Marcos Hernandez, Christope Herman, PJ Hastings and David Bates, all of BCM; Ben D Cox and LI-Ya Chiu, both of UT Austin; Elizabeth M luengas and Reuben S Harris of the University of Minnesota in Minneapolis; and Huong G Do and Lei Li, both of UT MD Anderson Cancer Center.