Proteome Analysis of Hordein-Null Barley Lines Reveals Storage Protein Synthesis and Compensation Mechanisms

Abstract

Hordeins are the major barley seed storage proteins and are elicitors of celiac disease. Attempts to reduce the hordein level in barley have been made; however, the resultant pleiotropic effects are less understood. Here, data-independent acquisition mass spectrometry was used to measure proteome-wide abundance differences between wild-type and single hordein-null barley lines. Using comparative quantitative proteomics, we detected proteome-wide changes (∼59%) as a result of the specific reduction in hordein proteins. The comparative analysis and functional annotation revealed an increase in non-gluten storage proteins, such as globulins and lipid transfer proteins, and proteins rich in essential amino acids in the null lines. This study yields an informative molecular portrait of the hordein-null lines and the underlying mechanisms of storage protein biosynthesis. This study indicates the extent to which protein content can be manipulated without biological consequence, and we envision its wide-scale application for studying modified crops.

Proteome and Nutritional Shifts Observed in Hordein Double-Mutant Barley Lines

“…To determine the magnitude of proteome perturbation, the mutant lines were compared with the WT ( Supplementary Figure 1 ). The proteins with perturbed abundance in the BC-mutant line was highly concordant with the changes observed in the parent B- and C-hordein mutant lines ( Bose et al, 2020 ). Additionally, the lys3 mutant, i.e.…”

“…A heatmap was generated from the differentially abundant proteins identified from pairwise comparisons between WT and DM lines to visualise the link between single and DM lines. The SM proteome abundance profiles were also included in the heatmap to allow contrast to the parent lines ( Figure 4 ) ( Bose et al, 2020 ). Hierarchical clustering shows the grouping of the biological replicates and that the D-hordein mutant line is closely linked to the WT, as D-hordein proportionately represents just 1–2% of total hordein content resulting in a minimal perturbation to the proteome.…”

“…Due to incomplete deletion of B-hordein locus, and the existence of a variety of hordein gene models mapped to the chromosome 1H ( Supplementary Figure 3B ), it is possible that the B-mutant line, Risø 56, carries additional hordein coding regions of which gene products were not detected herein. The remaining B-hordein-type proteins are evident in the B-mutant parent line where a hordein closely related to B-hordein subclasses was increased up to ∼8 fold ( Bose et al, 2020 ) as well as two low molecular weight B-hordeins (C7FB16, R9XWE6) from the B-hordein locus 2 in the BD-double mutant line ( Figure 5 and Supplementary Figure 3B ). Notably, the deletion of the major B-hordein locus did not affect the C-hordein locus.…”

Application of Mass Spectrometry-Based Proteomics to Barley Research

“…Implementing SWATH-MS, this study highlights one of the main applications of this advanced method in proteome-level alterations studies for investigating modified crops. 149 …”

Multi-Omics Strategies for Decoding Smoke-Assisted Germination Pathways and Seed Vigour

“…The advancement of mass spectrometry-based proteomics has enabled the routine measurements of thousands of proteins in terms of their abundance, localisation, interactions, and modifications while connecting the large-scale data to gene expression regulation and function [ 89 , 90 ]. Targeted and untargeted proteomics experiments have been widely used to investigate crop responses to the abiotic stress [ 89 , 90 , 91 , 92 ]. Proteomics studies have shown that plant-derived smoke water solutions can improve the abiotic stress tolerance capacity in soybean [ 3 ], chickpea [ 93 ] and rice [ 36 ].…”