Proteomics reveals commitment to germination in barley seeds is marked by loss of stress response proteins and mobilisation of nutrient reservoirs

Germination is a critical process in the reproduction and propagation of flowering plants, and is also the key stage of industrial grain malting. Germination commences when seeds are steeped in water, followed by degradation of the endosperm cell walls, enzymatic digestion of starch and proteins to provide nutrients for the growing plant, and emergence of the radicle from the seed. Dormancy is a state where seeds fail to germinate upon steeping, but which prevents inappropriate premature germination of the seeds before harvest from the field. This can result in inefficiencies in industrial malting. We used Sequential Window Acquisition of all THeoretical ions Mass Spectrometry (SWATH-MS) proteomics to measure changes in the barley seed proteome throughout germination. We found a large number of proteins involved in desiccation tolerance and germination inhibition rapidly decreased in abundance after imbibition. This was followed by a decrease in proteins involved in lipid, protein and nutrient reservoir storage, consistent with induction and activation of systems for nutrient mobilisation to provide nutrients to the growing embryo. Dormant seeds that failed to germinate showed substantial biochemical activity distinct from that of seeds undergoing germination, with differences in sulfur metabolic enzymes, endogenous alpha-amylase/trypsin inhibitors, and histone proteins. We verified our findings with analysis of germinating barley seeds from two commercial malting facilities, demonstrating that key features of the dynamic proteome of germinating barley seeds were conserved between laboratory and industrial scales. 1000 The results provide a more detailed understanding of the changes in the barley proteome during germination and give possible target proteins for testing or to inform selective breeding to enhance germination or control dormancy. Significance: Germination is critical to the reproduction and propagation of flowering plants, and in industrial malting. Dormancy, where seeds fail to germinate upon steeping, can result in inefficiencies in industrial malting. Our DIA/SWATH-MS proteomics analyses identified key changes during germination, including an initial loss of proteins involved in desiccation tolerance and germination inhibition, followed by decreases in lipid, protein and nutrient reservoir storage. These changes were consistent between laboratory and industrial malting scales, and therefore demonstrate the utility of laboratory-scale barley germination as a model system for industrial malt house processes. We also showed that dormant seeds that failed to germinate showed substantial biochemical activity distinct from that of seeds undergoing germination, consistent with dormancy being an actively regulated state. Our results provide a more detailed understanding of the changes in the barley proteome during germination and give possible target proteins for testing or to inform selective breeding to enhance germination or control dormancy.