Elucidation of translational regulatory mechanisms of plant immune responses

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Since plants do not have the specialized immune cells present in animals, activation of immunity in plants involves switching from producing growth-promoting proteins to defense proteins. Even though the process of generating templates (RNA transcripts) for defense proteins has been studied extensively, how the templates are selectively translated into proteins remains largely unknown. This major knowledge gap significantly impedes progress in developing new strategies of engineering disease resistant crops in agriculture as well as in bioengineering in general because controlled protein production is central to most biological functions. The long-term goal of this project is to elucidate the regulatory mechanisms by which plants produce defense proteins during immune responses. To achieve this goal, new technologies have been developed or adapted to examine the multiple steps connecting RNA transcripts to protein synthesis. Using translational switches to tighten the regulation of defense protein translation will minimize the yield penalty often associated with broad-spectrum disease resistance and revolutionize the practice of controlling crop diseases, which has mainly been focused on the disease-specific type of resistance. This project may also benefit other industries and medicine by improving recombinant protein production. The success of the project will also add to the pipeline of scientists from high school students to postdoctoral researchers with strong STEM training in performing frontier research. Identification of genes with altered translation efficiency during plant immune responses led to the discovery of novel regulators. Surprisingly, immune-related translation reprogramming in plants has been found to be distinct from the integrated stress response pathway found in yeast and mammals, indicating that plants use alternative regulatory mechanisms to control defense proteome translation. To elucidate these regulatory mechanisms, whole genome translatome analyses were performed for both pattern-triggered immune (PTI) and effector-triggered immunity (ETI) and a consensus sequence consisting of mostly purines (“R-motif”) was discovered in the 5’-leader sequences of translationally induced mRNAs during PTI. The R-motif was found to be not only necessary, but also sufficient for the PTI-induced translation. Its regulation is through the interaction with poly(A)-binding proteins (PABPs). Moreover, m6A-seq and SHAPE-MaP have been established to examine mRNA modifications and in vivo mRNA structural dynamics, respectively. With the establishment of these cutting-edge technologies, this project will focus on two specific aims: Aim 1: Study the effects of upstream open reading frames (uORF) on translation of the main ORF (mORF) in response PTI induction to test the hypothesis that RNA modification and secondary structure are critical determinants of ribosome accessibility to uORFs in controlling mORFs translation during PTI. Aim 2: Elucidate the signaling pathway leading to translatomic changes during PTI using both forward and reverse genetic approaches. The ultimate goal is to connect the various translational cis- and trans-elements with the currently known defense signaling network to fill the knowledge gap on how plants reprogram the proteome to mount immune responses.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.