In vivo, Quantitative label free proteomics and microscopy analysis highlights the central role of the endomembrane system during barley grain filling development.
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ABSTRACT: Abstract Barley is a cereal crop of worldwide importance for brewing industry, animal and in lesser extent humannutrition. Grain development is a tightly controlled process which predominantly contributes to the final grain quality. To understand molecular changes occurring in the early seed development stage up to the initial storage phase, proteome profiling of developing barley seed at 8, 10 18 and 20 days after flowering was analyzed. In vivo label free quantification approach allowed the quantification of more than 1200 proteins across the different stages. Confirmation of the reproducibility of the developmental data obtained in our proteomics analysis was performed using western-blot technics to validate the specific pattern of selected proteins and comparison with previously published data. Multi- and uni-variates statistics were used to unravel the biochemical dynamics underlying the grain filling process. Seed storage proteins (SSPs) were found to accumulate during the grain filling process suggesting a higher activity of the endoplasmic reticulum playing a major role in protein synthesis. This hypothesis was supported by a spatio-temporal analysis of the ER marker protein-disulfide-isomerase (PDI) at both transcript and protein level coupled with microscopic study of the ER dynamics. Results indicates that ER is predominantly active in the starchy endosperm up to 18 DAP and is subsequently degraded while the cells undergo program cell death. The central role of the ER was further strengthening by the analysis of all ER related proteins. Similarly, the pattern of vacuolar H+ ATPases suggested a decrease of the vacuolar acidity, microscopic analysis of the endosperm unravel a concomitant decrease of both vacuolar size and acidity associated with an abundance switch in different class of vacuolar annotated proteins (eg: Sucrose related metabolism proteins). Observed endomembrane system dynamics were supported by specific abundance changes of ESCRT related proteins such as VPS4 and SNF7 or VPS5 a protein associated to the retromer complex. Collectively, our results provide insight into endomembrane molecular processes taking place during barley early grain development, additionally the proteome dataset provide a comprehensive overview of a wide range of metabolic pathway which will be helpful for the cereal community.
INSTRUMENT(S): LTQ Orbitrap Elite
ORGANISM(S): Hordeum Vulgare (barley)
TISSUE(S): Seed
SUBMITTER: Valentin Roustan
LAB HEAD: Verena Ibl
PROVIDER: PXD009722 | Pride | 2018-07-02
REPOSITORIES: Pride
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