Mechanisms of light regulated translation in chloroplasts

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Energy from sunlight fuels life on earth through the process of photosynthesis. Light is both an essential resource and a source of stress, in that it damages the photosynthetic apparatus and other cellular structures through a process called photo-oxidative processes. As such, plants have evolved mechanisms that optimize the use of light, minimize light-induced damage, and promote repair of the photosynthetic apparatus should it occur. In this study the light-regulated synthesis of a gene product essential to this process, D1, will be examined. The rate of D1 synthesis changes rapidly in response to shifting light intensity and therefore the goal of this project is to elucidate the signaling pathways and mechanisms that mediate D1?s response. This problem will be addressed through genetic, molecular, and biochemical approaches in maize and Arabidopsis. These activities will provide research training for two undergraduate students, one PhD student, and a high school teacher. In addition, novel methods will be explored that have the potential to propel research on other topics. The hypotheses to be tested arise from recent ribosome profiling data that provide the first genome-wide views of the impact of light on translation in chloroplasts. Genetic, molecular, and biochemical approaches in maize and Arabidopsis will be employed, including a state-of-the-art method for identifying proteins bound to specific RNAs-of-interest in vivo, and a novel strategy involving designer RNA binding proteins. One set of experiments will test the hypothesis that the rapid light-induced change in ribosome occupancy on psbA mRNA (which encodes D1) is mediated by post-translational modification of proteins that bind the psbA 5' untranslated region. A second set of experiments will test the hypothesis that the global change in translation elongation rate is mediated by light-dependent post-translational modifications of proteins that stall ribosomes or catalyze translation elongation. A third set of experiments will address whether light induces these responses via the same signal transduction pathways that trigger other adaptations in response to photosynthetic activity. Finally, experiments will test the hypothesis that gene-specific translational regulators in chloroplasts, including those that regulate translation in response to light, generally act by modulating RNA structure proximal to ribosome binding sites.This project is co-funded by the Genetic Mechanisms Program in the Division of Molecular and Cellular Biosciences and by the Plant Genome Research Program in the Division of Integrative Organismal Systems.