Epigenetic Mechanisms for the Inheritance of Acquired Mutations

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This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5). RNA, normally thought of as a conduit in gene expression, has a novel mode of action in ciliated protozoa. The PI's lab has shown that maternal RNA templates provide both an organizing guide for DNA rearrangements in Oxytricha and a template that can transmit spontaneous mutations that may arise during somatic growth to the next generation. This opportunity for RNA-guided DNA repair is profound in the ciliate Oxytricha, which deletes 95% of its germline genome through global DNA rearrangements that severely fragment its chromosomes, and then sorts and reorders the hundreds of thousands of pieces remaining. The programmed information for reordering comes from transiently-expressed maternal RNAs. This means that changes that arise in the maternal somatic genome, whether at the level of point mutations or new DNA rearrangement patterns, have an opportunity to be passed on to the next generation, bypassing the usual mode of inheritance via germline DNA. Furthermore, the occasional transfer of point mutations in these RNA templates to the F1 generation provides a mechanism for stable inheritance of acquired, spontaneous somatic substitutions, without altering the germline genome. This suggests the somatic macronuclear genome is really an "epigenome", formed through templates and signals arising from the previous generation. The research will address 3 specific aims, which will yield detailed insight in Oxytricha into the following:1) Epigenetic inheritance of nucleotide substitutions via RNA templates, by exploring possible polymerases involved in RNA-guided DNA repair;2) Epigenetic inheritance of intron loss via programmed DNA deletion, which would contribute to developmental streamlining of a somatic genome;3) Epigenetic inheritance of copy-number variation: The third specific aim will explore an additional newly-discovered role for maternal template RNAs in regulating DNA copy number. This provides a heritable non-Mendelian mechanism for adaptation to the environment.Broader impacts: While the study of RNA templates and their expected development as novel synthetic tools to drive genome rearrangement are likely to have a great impact on the field of molecular biology, the first major impact of this research will be the scientific and professional training of three independent researchers (each will be the lead individual responsible for one of the three specific aims) in a broad new interdisciplinary area that combines RNA biology, DNA recombination, epigenetics, evolution, ciliate molecular biology, microbial diversity and genomics. All three specific aims address novel mechanisms for epigenetic inheritance, further developing Oxytricha as a model system. In addition to the training of doctoral and postdoctoral students, the investigator will also mentor undergraduates, an activity central to this project. All Princeton undergraduates perform intensive junior and senior independent work, often leading to publications, and the PI has a strong record of recruiting women and minorities to her lab and to Princeton's graduate program in Ecology and Evolutionary Biology. High school volunteers will also be invited to participate in research during the summer, and the opportunity to mentor younger students will provide the senior lab members with critical experience in training others to perform laboratory research. The investigator is also very active in developing a new model of publishing, as co-Editor-in-Chief of Biology Direct, which offers open, signed and published peer review and author responses, plus open-access readership, all of which encourage open scientific debate. The transparency of biology-direct.com also provides a novel vehicle for teaching students about the steps involved in publishing their work.