Dissecting the Trigger of the Chromatin Modification Cycle

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The living world functions through feed-forward and feed-back cycles, establishing self-perpetuating patterns that can persist over vast timescales. A major question that remains unanswered is how self-perpetuating biological cycles are first initiated. This project has the advantage of being able to trigger one such cycle reproducibly, generating a rare opportunity to understand how a biological cycle is originally initiated. This project uses cutting-edge techniques to understand the mechanism of the first trigger responsible for initiating a feed-forward cycle that regulates DNA newly introduced into plant cells. By placing new DNA into cells, this project will experimentally dissect the mechanism of the earliest stages initiating regulation. The results of this project will inform academics and industry on how to improve plants while avoiding the initiation of troublesome cycles of unwanted regulation. This project will also directly train the next generation of scientists, providing them with thorough mentoring to prepare them for careers in biology, data analysis and technology. In addition, this project will perform outreach to a new audience at the Jackie Joyner Kersee Center in East St. Louis. This program will benefit groups that have traditionally not seen themselves as eligible for a career in science due to lack of representative models in science, and is perfectly suited for the after school programs run by the Jackie Joyner Kersee Center. The establishment of chromatin modifications is an essential first step that leads to the long-term heritable epigenetic silencing of transposable elements and some transgenes. Once initiated, chromatin modifications such as cytosine DNA methylation, are replicated and, in plants, can be propagated at specific loci across generations for centuries. However, the mechanisms dictating how DNA methylation is first targeted to a locus is poorly understood. The preliminary data in support of this project involved building the tools to study the first round of new (de novo) chromatin modification, and these data demonstrate that the production of scaffolding, non-coding RNAs at a locus is the key, milestone event required for de novo DNA methylation. This kind of event can be best studied in plants due to the recruitment of a specific RNA Polymerase (Pol V) to generate these scaffold RNAs. The overarching goals of this project are to understand how scaffold RNA target regions are selected and how Pol V is first recruited during de novo DNA methylation. This work utilizes cutting-edge techniques to dissect, at the molecular level, an unknown mechanism responsible for establishing de novo DNA methylation and triggering the long-term biological cycle of heritable, epigenetic silencing of transposable elements and transgenes.This project is co-funded by the Genetic Mechanisms Program in the Molecular and Cellular Biosciences Division and by the Plant Genome Research Program in the Integrative Organismal Systems Division.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.