Investigator: Alexandra Deaconescu, Brown University

Mentor: Rebecca Page, Brown University

Scientific Theme: Cancer and Molecular Toxicology

Abstract: In the last decade, ATP-dependent dsDNA translocases remodeling protein-nucleic acid assemblies have emerged as key players for cellular physiology, having been implicated in pervasive diseases such as cancer, diabetes as well as neurodegeneration and accelerated aging. A case in point is the large family of ATP-dependent transcription-repair coupling factors (also known as Mfd in bacteria and CSB in humans) that recognize and remodel/dissociate transcription complexes stalled at DNA lesions, mobilize nucleosomes (in the human system) and recruit nucleotide excision repair enzymes to promote strand-specific repair of DNA lesions in the template strand. The activity crucial for the function of these proteins is their ability to translocate directionally on dsDNA in a highly regulated manner. However, we currently have a poor understanding of their mechanochemical cycle and DNA binding/translocation strategy, primarily due to the scarcity of structural information on these proteins. Transcription-repair coupling factors are akin to the intensely studied chromatin remodeling factors, yet structural knowledge of chromatin remodeling factors cannot easily be extended to Mfd and CSB due to the unique architecture of these proteins, with family- specific regulatory domains that inhibit the ATP-dependent core. Our overall goal is to dissect at atomic resolution the mechanism of dsDNA translocation utilized by transcription-repair coupling factors and elucidate the conformational changes that these proteins undergo for activation upon binding to their targets. To this end, we will use a complementary approach combining biochemical and biophysical techniques with structural determination methods such as by X-ray crystallography and single-particle electron microscopy. Our studies will provide atomic models for key intermediates in the pathway, shedding light on how transcription-coupled repair is initiated and regulated, enabling a mechanistic understanding of disease mutations associated with transcription-repair coupling factors, and serving as a springboard for more targeted experimentation in cells and animal models, and ultimately, improved therapies.

Human Health Relevance: Transcription-coupled DNA repair has long been associated with Cockayne Syndrome, an accelerated aging disease that manifests itself through severe neurodegeneration and developmental defects, but also, more recently, with several types of cancer. In bacteria, transcription-repair coupling factors are important for the development of resistance to floroquinolones. Thus, our studies are relevant for the development of new antimicrobials and alternative methods of chemotherapy.