Bioluminescence DNA Repair Reporter To Detect DNA Break Repair
Researchers from Massachusetts General Hospital (MGH) and the Academia Sinica in Taiwan have developed a new bioluminescent reporter that tracks DNA double-stranded break (DSB) repair in cells. DNA repair pathways can be monitored directly in animals as well as cell lines using the novel bioluminescent repair reporter (BLRR)-based system developed by the international team. For in vivo studies, no such system existed previously. In multiple conditions, including cancer, these pathways play a crucial role.
The co-senior author of the paper, an investigator in the Department of Neurology at MGH, Christian Elias Badr, Ph.D., explains, “Cells can inherently repair the DNA damage caused by radiation and chemotherapy, and this is one of the main reasons cancer cells are resistant to treatment.” Charles Pin-Kuang Lai Ph.D., the Academia Sinica in Taiwan is the study’s other co-senior author. In the Nucleic Acids Research, their study appeared as an online advance paper.
The key to maintaining cell viability and genomic integrity is DSB damage repair. It also plays a role in cancer treatment, as DSB is disrupted by chemoradiotherapy (radiation and chemotherapy) involved in cancer treatments. To reduce cell death caused by DSB, a cell
can recognize the damage and can use its intrinsic DNA damage response (DDR). As a result, drug resistance and recurrence in some malignancies can be promoted by the cancer cell’s own DNA repair mechanisms.The members of the team had worked on enzymes called luciferases, and the BLRR approach builds on this work. Luciferases are useful for tracking molecules in cells as these produce bioluminescence. To detect the two major pathways to DSB repair— homology-directed repair (HDR) and non-homologous end joining (NHEJ), BLRR uses secreted Gaussia and Vargula luciferases. The HDR and NHEJ-related activities can be tracked over time in cells by the researchers and it also detects DSB repairs in xenografted tumors in vivo.
Badr says, “DNA damage in cells can be studied with next-generation sequencing (NGS), however, it is more time-consuming and costly. The accuracy of our system is comparable to NGS.”
Multiple studies were carried out by the researchers using their new tag. A notable difference in the efficiency of CRISPR/Cas9-mediated editing with guide RNAs 1-10bp apart was found in one of the studies. Also, altered dynamics for DSB repair induced by small-molecule modulators were detected using BLRR analysis. Additionally, by inhibiting DNA repair protein RAD51 homolog 1, they used the system to discover HDR-suppressing functions of anticancer cardiac glycosides in human glioblastomas and glioma cancer stem-like cells.
According to the authors, the BLRR system was described in their paper as a highly sensitive platform to longitudinally and simultaneously track HDR and NHEJ dynamics that is sufficiently versatile for interpreting the DSB repair’s physiology and therapeutic development. To identify novel therapeutics that sensitize cancer cells to radiation and chemotherapy, the authors plan on using this reporter system in high throughput drug screening.
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Bioluminescence DNA Repair Reporter To Detect DNA Break Repair Created By Researchers
