Poster Presentation HUPO 2019 - 18th Human Proteome Organization World Congress

Understanding the storage protein biosynthesis and protein compensation in hordein double-null barley lines using SWATH mass spectrometry (#439)

Utpal Bose 1 , James A. Broadbent 1 , Keren Byrne 1 , Malcolm Blundell 2 , Crispin A. Howitt 2 , Michelle L. Colgrave 1
  1. Agriculture and Food, CSIRO, St Lucia, QLD, Australia
  2. Agriculture and Food, CSIRO, Canberra, ACT, Australia


The hordeins are the major barley seed storage proteins and are elicitors of coeliac disease - a condition that affects ~1% of the world population. There are four multi-genic hordein families in barley: B-hordeins representing 70-80% of total gluten; C-hordeins (10-20%); D-hordein (<5%); and, γ-hordeins (1-2%). Using conventional breeding a series of null lines were developed in which different classes or combinations of classes of hordeins are absent.  


Data-independent acquisition (DIA) mass spectrometry (MS) was used to measure proteome-wide abundance differences between wild-type and selectively-bred hordein double-null barley lines. Data were acquired in information-dependent acquisition (IDA) and sequential window acquisition of all theoretical fragment-ion spectra-mass spectrometry (SWATH-MS) on a TripleTOF 6600 MS (SCIEX, USA). Statistical and functional analyses were performed on the proteins perturbed between wild-type and hordein double-null barley lines.


A total of 6,138 peptides mapping to 1,907 proteins were quantified at a 1% false discovery rate. Pairwise comparisons revealed proteome-wide alterations for BC-, BD- and CD-null lines in the order of ~16%, ~10% and ~14%, respectively. As an example, the comparison between wild-type and the BC-null line identified 151 up-regulated proteins (7.9%), while 145 (7.6%) proteins were down-regulated in the BC-null line. Contextualization within a protein-protein interaction network reveals the up-regulation of proteins associated with primary metabolism, transcription and enzymatic biosynthesis processes while down-regulation of heat shock proteins can be found in the BC-null line. Gene Ontology (GO) analysis (molecular function) revealed that enzymatic activities were up-regulated, whilst nutrient reservoir activities were down-regulated in the BC-null line.

Concluding remarks

Overall, the GO-based analysis provides an overview of the functional classes that are perturbed during the breeding process. Grain proteome profiling delivers an informative molecular portrait of the hordein double-null lines and the underlying storage protein biosynthesis mechanisms thereby shedding light on mechanism of proteome compensation.