Most eukaryotic cells contain mitochondria, the membrane-bound organelles that produce energy for the cell. In addition to DNA found in the nucleus of the cell, mitochondria contain DNA. The faithful maintenance of the mitochondrial DNA is essential to the normal function of cells. Deletions of large segments of the mitochondrial DNA are common mutations in many types of cells. In humans, these mutations accumulate with age and are associated with inherited disease syndromes. Deletions of this type are also commonly observed in the mitochondrial DNA of baker's yeast. The long-term goal of this project is to understand the mechanism by which these deletions arise using yeast as a model system. The researchers aims are directed at understanding the proteins involved in this process and their activities. The researchers have devised genetic strategies to study these factors to study the proteins that contribute to mitochondrial deletions, determine what proteins bind to double-strand breaks in mitochondrial DNA, and study the temporal events in mitochondrial double-strand break repair. Completion of this work will provide new information on the broad area of mitochondrial DNA metabolism, including replication, recombination and repair.
Broader impacts:
This work will have a positive impact in the education of undergraduates in the biological sciences in several ways. In the past 5 years, the principal investigator has mentored 17 undergraduate students pursuing independent projects. Studies with yeast are well suited to undergraduates with little experience, allowing them to learn a large number of techniques and concepts in a summer or semester. In addition, the research represents a collaboration with the co-principal investigator at SUNY-Brockport, which is primarily a 4-year institution that does not grant Ph.D. degrees. The Department of Biological Sciences provides excellent opportunities for undergraduate research. More to the point, the record of the co-principal investigator over the past 5 years has supported the goal of the Department of Biological Sciences at Brockport to further undergraduate involvement in research. The co-principal investigator has sponsored a total of 15 undergraduates, and 7 Master's students in a variety of independent studies and directed research projects, 3 of those were CSTEP/McNair Scholars. The researchers will continue to honor this commitment to scientific education with the involvement of undergraduate biology majors at SUNY Brockport and the University of Rochester in the current project. Both the principal investigator and the co-principal investigator have adapted several small projects for use in laboratory courses. Exercises have been designed that incorporate standard yeast genetic approaches and yeast two-hybrid techniques to teaching labs. The students perform screens with baits relevant to studies currently being performed in both the principal investigator's and the co-principal investigator's laboratories.
Humans, and the much simpler yeast are eukaryotes. Eukaryotic cells contain organelles that are separated from the rest of the intracellular material by membranes. Among these organelles are mitochondria, whose function is to produce energy for the cell. Like the nucleus of the cell, mitochondria contain DNA genomes, and their maintenance without error is essential to the normal function of cells. Mutations to mitochondrial DNA often impair energy production, causing global cellular dysfunction. Deletions of large segments of the mitochondrial DNA are common mutations in many types of cells. In humans, these mutations accumulate with age and are associated with inherited disease syndromes. Deletions of this type are also commonly observed in the mitochondrial DNA of baker’s yeast. Very important cellular processes, including those required to maintain the nuclear genome, are similar in nearly all eukaryotes, so studies performed in yeast have provided significant insight into human processes. The molecular tools available for research in yeast have made it a powerful tool for the study of mitochondrial biology as well. The long-term goal is to understand the fundamental cellular processes that contribute to mitochondrial deletion mutations. The studies performed to date have revealed that many of the proteins known to be involved in similar events in the nuclear genome generate deletions in mitochondria, however, the contributions of the proteins to deletions in each cellular compartment are not the same. Because deletions in the nucleus are generated by aberrant repair of breaks to both strands of a DNA double helix, mitochondrial breaks were also investigated. This required the engineering of a system to generate a specific, controlled, repairable double-strand break in the mitochondrial genome. With this powerful new tool, additional proteins have been identified that are important for mitochondrial DNA repair and the generation of deletions, that would not otherwise have been identified. Many of these yeast proteins have human counterparts, suggesting that these pathways may also be important in the repair of human mitochondrial DNA.
