Nutrient remobilization from senescing tissues to developing grains is a tightly regulated process that affects the nutritional value of crops. We have recently shown the existence of a close connection between senescence and nutrient remobilization through the cloning of the wheat GPC (Grain Protein Content) gene. RNA interference (GPC-RNAi) plants showed delayed senescence and reduced grain mineral content by over 30%. The GPC gene is a NAC transcription factor involved in the early regulation of leaf-senescence and mineral remobilization to the grains. We propose to take a first step towards unraveling these processes by using GPC as an early entry-point to understand the regulatory network controlling senescence and consequently, the initial stages of nutrient remobilization. Our first objective is to study the individual contribution of the paralogous GPC-1 and GPC-2 genes to senescence and nutrient remobilization in wheat by field experiments. To accomplish this objective we have developed ethyl methane sulphonate (EMS) mutant population and identified knockout mutants for each copy of these two genes. We hypothesize that these mutants will show delayed senescence ('ever-green') phenotype and might have an effect on grain yield. Our second objective is to identify and study the genes regulated by GPC-1 and GPC-2. We will validate a list of GPC-regulated genes previously identified by comparing the transcriptome of GPC-RNAi transgenic and its non-transgenic control line through massively parallel next generation sequencing (NGS) experiments. Selected GPC-regulated genes will be validated by quantitative PCR in the GPC mutants and transgenic wheat over-expressing GPC-1. The most promising candidates will be further investigated by generating new EMS mutants. The experiments proposed here will increase our understanding of the gene regulatory network that governs senescence and nutrient remobilization in wheat and the specific roles of the two GPC paralogues.
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