Abstract
Cereal endosperm is a short-lived tissue adapted for nutrient storage, containing specialized organelles, such as protein bodies (PBs) and protein storage vacuoles (PSVs), for the accumulation of storage proteins. During development, protein trafficking and storage require an extensive reorganization of the endomembrane system. Consequently, endomembrane-modifying proteins will influence the final grain quality and yield. However, little is known about the molecular mechanism underlying endomembrane system remodeling during barley grain development. By using label-free quantitative proteomics profiling, we quantified 1,822 proteins across developing barley grains. Based on proteome annotation and a homology search, 94 proteins associated with the endomembrane system were identified that exhibited significant changes in abundance during grain development. Clustering analysis allowed characterization of three different development phases; notably, integration of proteomics data with in situ subcellular microscopic analyses showed a high abundance of cytoskeleton proteins associated with acidified PBs at the early development stages. Moreover, endosomal sorting complex required for transport (ESCRT)-related proteins and their transcripts are most abundant at early and mid-development. Specifically, multivesicular bodies (MVBs), and the ESCRT-III HvSNF7 proteins are associated with PBs during barley endosperm development. Together our data identified promising targets to be genetically engineered to modulate seed storage protein accumulation that have a growing role in health and nutritional issues.
Highlights
Cereal endosperm is a short-lived tissue adapted for nutrient storage, containing specialized organelles, such as protein bodies (PBs) and protein storage vacuoles (PSVs), for the accumulation of storage proteins
The seed weight between the stages are significantly different except the stage between 6 and 10 days after pollination (DAP) (Supplementary Fig. S2e) These results are in line with the previously published microscopic data where the most dramatic endomembrane rearrangements could be observed between 8 and 12 DAP13
Proteomics and in situ microscopic analyses enable the mapping of the endomembrane system of developing barley endosperm
Summary
Cereal endosperm is a short-lived tissue adapted for nutrient storage, containing specialized organelles, such as protein bodies (PBs) and protein storage vacuoles (PSVs), for the accumulation of storage proteins. Endomembrane-modifying proteins will influence the final grain quality and yield. Multivesicular bodies (MVBs), and the ESCRT-III HvSNF7 proteins are associated with PBs during barley endosperm development. The starchy endosperm thereby is characterized as a storage site, accumulating starch and seed storage proteins (SSPs)[2]. SSPs, which account for more than 50% of the grain protein content[3,4], accumulate in the outer layer of the endosperm, in the subaleurone, and in the starchy endosperm, the latter in parallel with starch granules[5]. The SSP trafficking routes depend on the cereal species, endosperm layer and development stage[7,8,9]. Endomembrane-modifying proteins within the endomembrane system will have an influence on the final grain quality/yield and recombinant protein production
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