The cell wall is an important compartment in grain cells that fulfills both structural and functional roles. It has a dynamic structure that is constantly modified during development and in response to biotic and abiotic stresses. Non-structural cell wall proteins (CWPs) are key players in the remodeling of the cell wall during events that punctuate the plant life. Here, a subcellular and quantitative proteomic approach was carried out to identify CWPs possibly involved in changes in cell wall metabolism at two key stages of wheat grain development: the end of the cellularization step and the beginning of storage accumulation. Endosperm and outer layers of wheat grain were analyzed separately as they have different origins (maternal and seed) and functions in grains. Altogether, 734 proteins with predicted signal peptides were identified (CWPs). Functional annotation of CWPs pointed out a large number of proteins potentially involved in cell wall polysaccharide remodeling. In the grain outer layers, numerous proteins involved in cutin formation or lignin polymerization were found, while an unexpected abundance of proteins annotated as plant invertase/pectin methyl esterase inhibitors were identified in the endosperm. In addition, numerous CWPs were accumulating in the endosperm at the grain filling stage, thus revealing strong metabolic activities in the cell wall during endosperm cell differentiation, while protein accumulation was more intense at the earlier stage of development in outer layers. Altogether, our work gives important information on cell wall metabolism during early grain development in both parts of the grain, namely the endosperm and outer layers. The wheat cell wall proteome is the largest cell wall proteome of a monocot species found so far.

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Seeds of the model grass Brachypodium distachyon are unusual because they contain very little starch and high levels of mixed-linkage glucan (MLG) accumulated in thick cell walls. It was suggested that MLG might supplement starch as a storage carbohydrate and may be mobilised during germination. In this work, we observed massive degradation of MLG during germination in both endosperm and nucellar epidermis. The enzymes responsible for the MLG degradation were identified in germinated grains and characterized using heterologous expression. By using mutants targeting MLG biosynthesis genes, we showed that the expression level of genes coding for MLG and starch-degrading enzymes was modified in the germinated grains of knocked-out cslf6 mutants depleted in MLG but with higher starch content. Our results suggest a substrate-dependent regulation of the storage sugars during germination. These overall results demonstrated the function of MLG as the main carbohydrate source during germination of Brachypodium grain. More astonishingly, cslf6 Brachypodium mutants are able to adapt their metabolism to the lack of MLG by modifying the energy source for germination and the expression of genes dedicated for its use.

With the increase in the use of plant ingredients in food and the growing demand for pulse protein ingredients, dry fractionation is a sustainable method for producing high-quality protein fractions from pulses. The objective of this work was to assess the morphology of pea fractions obtained by dry fractionation, and their detailed composition with a focus on non-protein components, especially lipids. Pea seeds from a single cultivar were first dehulled, then milled via either stirred ball milling or impact milling, which affected the microstructure of the flours. For instance, scanning electron microscopy revealed that starch granules looked damaged after stirred ball milling, whereas impact milling led to flours with smooth starch granules. Air classification led to a protein-rich fraction (concentrate) with at least 46 g protein /100 g dry matter (d.m.) and to a protein-poor fraction with maximum 18 g protein /100 g (d.m.). The lipids endogenously present in pea seeds (around 3 g/100 g (d.m.) in the starting flour) accumulated in the protein-rich fractions (> 5 g/100 g (d.m.), at the expense of the protein-poor fractions where the lipid content decreased compared to the flour. The same trend was obtained for the ash content. Finally, protein solubility in the flours and protein-rich fractions was investigated in aqueous suspensions at neutral pH, and could be improved by applying a high-pressure homogenization treatment. The microstructure of the suspensions (non-treated, or treated with high-pressure homogenization) was also characterized by confocal laser scanning microscopy. These outcomes expand the current knowledge of dry-fractionated pulse protein concentrates, which is particularly important when targeting their rational use for food applications.