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Using advanced imaging technology, scientists at the Mayo Clinic have provided unprecedented understanding of the BRCA1-BARD1 protein complex, which is often mutated in patients with breast or ovarian cancer. Their article, published in Nature, identifies aspects of how BRCA1-BARD1 works, supporting future translational research, cancer prevention efforts and drug development.
“BRCA1-BARD1 is important for DNA repair. It is directly related to cancer because hundreds of mutations in the BRCA1 and BARD1 genes have been identified in cancer patients, ”says Georges Mer, Ph.D., structural biologist and biochemist at the Mayo Clinic. who is the main author of the article. “But no one knows whether these mutations, or variants of unknown importance, predispose to cancer or not because we don’t know if the variants are located in a region of BRCA1-BARD1 that is important for function. Now because we can see how BRCA1-BARD1 works, we have a good idea of which regions of BRCA1-BARD1 are important for function. “
In a cell, the complex of DNA and histone proteins is complexed into what is called chromatin and packaged in bundles called nucleosomes. DNA damage response proteins must access chromatin to repair damaged DNA. BRCA1-BARD1 contributes to the attachment of broken DNA strands, which aids in the maintenance and survival of cells. But it’s also a function that could eventually be blocked or inactivated if this is a strategy a cancer cell uses to survive chemotherapy.
Cryoelectron microscopy and nuclear magnetic resonance spectroscopy
“We used two techniques – electron cryomicroscopy and nuclear magnetic resonance spectroscopy – to understand at near atomic resolution how BRCA1-BARD1 associates with the nucleosome, the repeating unit of chromatin, and how BRCA1-BARD1 alters the chromatin, ”says Dr Mer.
In electron cryomicroscopy, purified BRCA1-BARD1 bound to nucleosomes, called together macromolecules, are frozen and then imaged using an electron microscope. Macromolecules are oriented in various ways within the sample, so a computer program evaluates all of the orientation data to create a 3D structure. Dr Mer and his team also examined BRCA1-BARD1 nucleosome complexes with nuclear magnetic resonance spectroscopy, which uses a strong magnet to probe the relative positions of atoms in macromolecules. Using these imaging tools, scientists were able to visualize BRCA1-BARD1 in action and discover a new function of the complex.
“We showed how BRCA1-BARD1 attaches ubiquitin to the nucleosome, but we also determined that BRCA1-BARD1 recognizes ubiquitin already attached to the nucleosome, which serves as a signal for broken DNA,” says Dr. Mer. We have discovered an unexpected interference whereby recognition of ubiquitin by BRCA1-BARD1 enhances its ubiquitin binding activity, and this helps us better understand how BRCA1-BARD1 performs its function. “
The researchers created a video from the electron cryomicroscopy data to show where the protein complex interacts with the nucleosome [see link below].
From discovery science to patient care
Dr. Mer and his team expect that the high-resolution images of BRCA1-BARD1 can help guide patient care and future cancer treatment in two ways: by classifying variants of unknown importance and by orienting the focus. drug development with more precision.
“With these 3D structures, we should be able to convert several variants of unknown importance into variants that may predispose to cancer,” says Dr Mer. “This work should also have an impact on long-term drug development because the 3D structures of BRCA1-BARD1 in complex with the nucleosome we have generated can aid in the design of small molecules that could, for example, inactivate BRCA1-BARD1. “
In addition to Dr Mer, the other authors of the article are Qi Hu, Ph.D .; Maria Victoria Botuyan, Ph.D .; Debiao Zhao, Ph.D.; Gaofeng Cui, Ph.D.; and Elie Mer. This research was funded by the National Institutes of Health, the Mayo Clinic Cancer Center, the Mayo Clinic Center for Biomedical Discovery and the Ovarian Cancer Research Alliance, and was made possible through cryo-electron microscopy and at the nuclear magnetic resonance instrumentation of the Pacific Northwest Center for Cryo-EM and Mayo Clinic, respectively.
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Material provided by Mayo Clinic. Original written by Sara Tiner. Note: Content can be changed for style and length.
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