Role of ISWI Chromatin Remodeler in Transcription-coupled Repair and Genome Stability

Tufts University, Medford MA

Investigators

Abstract

Intellectual Merit. Cells of every organism must preserve the genome to prevent mutations. Repetitive DNA is common in genomes and presents a particular challenge for cells, as slippage can occur during replication or repair. In particular, structure-forming repeats, such as CAG/CTG triplet repeats, have been shown to be particularly prone to gains and losses in copy number. It is essential to understand mutational processes within repetitive DNA, especially when mutations occur in a region that could affect gene function. One of the key repair systems used within transcribed regions is transcription-coupled repair. Recent discovery indicates that a new factor, the Isw1 chromatin remodeler protein, is needed to prevent CAG expansions during transcription-coupled repair. The specific objective of this project is to determine the role of Isw1 in transcription-coupled repair, and how it controls fidelity of repair within a CAG repeat to prevent expansions. The model eukaryote Saccharomyces cerevisiae (baker's yeast) will be used since it exhibits repeat instability similar to human cells, yet offers powerful genetic and molecular tools that will be employed to address the proposed questions. This research will provide fundamental insight into mutational processes in eukaryotic organisms, including yeast and humans. In addition, the discovery of a new player in transcription-coupled repair, the Isw1 protein, creates an opportunity to better understand this important DNA repair pathway and how it operates in the context of chromatin, which is the form that DNA takes when packaged within cells. Isw1 can "remodel" chromatin, or change its structure to facilitate access to the DNA. Thus the results of this project also have a high potential to significantly advance the current limited knowledge of the role of chromatin remodeling in DNA repair. Broader Impacts. In addition to creating new scientific knowledge, this project will have a broader impact on education of students at the undergraduate level. Traditionally laboratory exercises taught in undergraduate courses consist of activities where students follow a set protocol with a particular result in mind. However, real science rarely follows this predictable pattern. Part of this project will be to take advantage of the versatility of our model system to develop a new laboratory course designed to engage Tufts undergraduates in novel research in the area of molecular genetics. This new course will allow undergraduates to experience "non-cookbook" laboratory research and make new discoveries by doing an open-ended screen for cellular factors that protect against mutations and genome rearrangements. Currently this opportunity is available to only a small number of students, so the availability of this laboratory will significantly broaden the number of students engaged in significant scientific research and critical thinking. The young scientists working on the transcription-coupled repair project will also be involved in the design and implementation of the new course, giving them the opportunity to combine their research efforts with course design and training in teaching. To allow other educators to utilize these teaching modules, once finalized, the course materials, protocols and reagents will be made available to the larger Genetics community through publication.