Proteomic analysis of proteins responsive to drought and low temperature stress in a hard red spring wheat cultivar

Drought stress is becoming more prevalent with global warming, and has been shown to have large effects on gluten proteins linked to wheat bread making quality. Likewise, low temperature stress can detrimentally affect proteins in wheat. This study was done to determine the differential expression of high molecular weight (HMW) gluten proteins in a drought and low temperature stressed high quality hard red spring wheat cultivar (PAN3478), against a control. The three treatments were applied in the greenhouse when the main tillers of each pot were at the soft dough stage, on 15 pots per replication, three replications and three plants per pot for each treatment. Seed of main tillers were bulked and randomly sampled. Removal of gliadins was done with 50% (v/v) 1-propanol. The HMW proteins were then extracted and separated by 2-dimensional gel electrophoresis. Gels were analyzed with SameSpots Progenesis (version 4.6.1.218). The protein spots that had p values lower than 0.05 and fold value equal to or greater than 1.2 were considered significantly differentially expressed. Spots were excised from the gel, digested with trypsin and analysed by mass spectrometry. The proteins were identified by matching the spectra to theoretical data from the Swissprot protein database. There was a 1.3 to 1.8 fold change in 19 protein spots due to the cold treatment. The drought treatment caused 1.3 to 3.8 fold change in 20 spots that were significantly differentially expressed. Two spots, one with matching peptide sequences to HMW glutenin proteins DX5, DY10 and PC256, and one to HMW glutenin protein PW212, were differentially expressed under both drought and cold stress. Other proteins differentially expressed under cold stress included proteins with similar peptides to alpha/beta-gliadin A-IV, fructan 1- exohydrolase w3, a number of HMW-GS P212 and HMW-GS DY12 proteins and HMW-GS DY10, chloroplastic Nad(P)H-quinone oxidoreductase subunit 2 A, alpha/beta-gliadin and low molecular weight-GS PTDUCD1. Under drought stress ATPase, HMW-GS PW212, HMW/GS (HMW subunit 12) and DY10, Ubiquitin and Eukaryotic translation initiation factor isoform 4G-2 were differentially expressed. The HMW glutenins are known to have a large effect on baking quality, and clearly protein spots with similar peptide sequences with especially HMW-GS DY12, PW212, PC256, HMWGS DX5 and DY10 were responsive to cold and drought stress, and could directly influence baking quality.