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Acta Agronomica Sinica ›› 2021, Vol. 47 ›› Issue (7): 1248-1258.doi: 10.3724/SP.J.1006.2021.01062

• CROP GENETICS & BREEDING · GERMPLASM RESOURCES · MOLECULAR GENETICS • Previous Articles     Next Articles

Differentially expressed protein analysis of different drought tolerance hulless barley leaves

LI Jie, FU Hui, YAO Xiao-Hua, WU Kun-Lun*()   

  1. Academy of Agricultural and Forestry Sciences, Qinghai University / Qinghai Academy of Agricultural and Forestry Sciences / Qinghai Key Laboratory of Hulless Barley Genetics and Breeding / Qinghai Subcenter of National Hulless Barley Improvement, Xining 810016, Qinghai, China
  • Received:2020-08-05 Accepted:2020-12-01 Online:2021-07-12 Published:2021-03-14
  • Contact: WU Kun-Lun E-mail:wklqaaf@163.com
  • Supported by:
    This study was supported by the National Natural Science Foundation of China(32060480);This study was supported by the National Natural Science Foundation of China(31660388);This study was supported by the National Natural Science Foundation of China(31960427);the Qinghai Provincial Academy of Agriculture and Forestry Innovation Fund(2018-NKY-12);the China Agriculture Research System(CARS-05);the Qinghai Science and Technology Support Project(2019-ZJ-7075)

Abstract:

In order to reveal the differences in response to drought stress among different hulless barley varieties from the protein level and to analyze the protein molecular mechanism of drought tolerance, Handizi Barley (HDZ) resistant to drought stress and Dama Barley (DM) sensitive to drought stress were used as research materials in this study. Drought treatment were determined by potted-planting method with limited water supply, four physiological indexes of hulless barley leaves with different drought gradients, including chlorophyll, soluble protein, malondialdehyde content, and relative conductivity were investigated. iTRAQ technology was used to conduct differential protein analysis on the whole protein group of barley leaves under deep drought stress. The results showed that with the extension of the drought treatment, the chlorophyll and soluble protein content of two hulless barleys under drought stress gradually decreased, the electrical conductivity and malondialdehyde content gradually increased, and the decrease in chlorophyll and soluble protein content, the increase in electrical conductivity and the content of malondialdehyde in Dama were greater than that of Handizi; 4163 proteins (polypeptides) were quantified, among them, compared with normal culture in the Handizi comparison group, 68 up-regulated proteins and 63 down-regulated proteins were screened by iTRAQ; in the comparative group of Dama, 21 up-regulated proteins and 32 down-regulated proteins were screened. KEGG pathway showed that the top three enrichment pathways were metabolic, amino acid biosynthesis, and secondary metabolite biosynthesis. The first one mainly related to citric acid cycle, carbon cycle, and other metabolic pathways. The synthesis and degradation of amino acids were mainly involved arginine and alanine. The synthesis of secondary metabolites were about arachidonic acid and linolenic acid. This study screened the proteins related to the metabolic pathways and other related functions in response to drought stress on proteome level in hulless barley, providing a theoretical basis for revealing the molecular regulation mechanism in response to drought stress.

Key words: hulless barley, drought stress, leaf, differentially expressed protein, iTRAQ

Fig. 1

Chlorophyll content of HDZD-HDZCK and DMD- DMCK A: chlorophyll a; B: chlorophyll b; C: total chlorophyll. Uppercase letters indicate extremely significant differences at the 0.01 probability level, lowercase letters indicate significant differences at the 0.05 probability level."

Table 1

Effects of drought treatment on protein, relative conductivity and malondialdehyde content of barley leaves"

处理
Treatment
品种
Variety
蛋白质含量
Protein content
(mg g-1)
相对电导率
Relative electrical
conductivity (%)
丙二醛含量
Malondialdehyde content
(μmoL g-1)
对照
CK
旱地紫 HDZ 7.30±0.10 Aa 44.50±0.68 Cc 32.92±0.45 Cc
大麻 DM 10.00±0.56 Aa 50.10±0.34 Cc 28.70±0.10 Cc
干旱处理7 d
7 days after drought treatment
旱地紫 HDZ 6.90±0.55 Aa 45.89±0.47 Bb 35.55±0.29 Bb
大麻 DM 7.87±0.12 Bb 54.33±0.60 Bb 32.21±0.38 Bb
干旱处理10 d
10 days after drought treatment
旱地紫 HDZ 6.75±0.26 Aa 48.97±0.71 Aa 38.92±0.42 Aa
大麻 DM 7.65±0.61 Bb 60.22±0.39 Aa 40.33±0.48 Aa

Table 2

Distribution of coverage percentage of identified protein"

比较组
Comparison group
肽段覆盖率 Protein coverage (%) 总计
Total
≤10 10<X≤20 20<X≤30 30<X≤40 40<X≤50 50<X≤60 60<X≤70
HDZD-HDZCK 650 807 409 192 64 18 8 2150
DMD-DMCK 622 752 390 170 55 16 6 2013

Fig. 2

Differentially expressed protein volcano map of HDZD-HDZCK barley (left) and DMD-DMCK barley (right)"

Fig. 3

GO enrichment of differentially expressed proteins A: biological process (BP), B: cell component (cc), and C-G: molecular functions (MF). A: defense response; B: extracellular part; C: catalytic activity; D: ATP binding; E: structural molecule activity; F: transfer activity; G: transport activity; H: other activities."

Table 3

Functional classification of differentially expressed proteins of HDZD-HDZCK and DMD-DMCK"

蛋白质功能分类
Functional classification of proteins
蛋白质数量
Number of proteins
胁迫应答类 Stress response 11
脂类代谢相关类 Lipid metabolism 13
糖类代谢相关类 Carbohydrate metabolism 10
蛋白质代谢相关类 Protein metabolism 48
氧化还原类 Oxidoreduction 35
能量供应类 Energy 16
光合相关类 Photosynthetic 15
未知蛋白 Unknown functional protein 36

Fig. 4

KEGG annotation of differentially expressed proteins of HDZD-HDZCK (left) and DMD-DMCK (right)"

Table 4

KEGG pathway of differentially expressed proteins"

通路ID
Pathway ID
通路名称
Pathway name
ko00020 三羧酸循环 TCA cycle
ko00640 丙酸酯代谢 Propanoate metabolism
ko01200 碳代谢 Carbon metabolism
ko00270 半胱氨酸和蛋氨酸代谢 Cysteine and methionine metabolism
ko00620 丙酮酸代谢 Pyruvate metabolism
ko00630 乙醛酸和二羧酸的代谢 Glyoxylate and dicarboxylate metabolism
ko00710 光合作用中的碳固定 Carbon fixation in photosynthetic
ko00195 光合作用 Photosynthsis
ko03010 核糖体 Ribosome
ko03013 RNA转运 RNA transport
ko00190 氧化磷酸化 Oxidaxtive phosphorylation
ko04145 吞噬作用 Phagosome
ko04141 内质网的蛋白质加工 Protein processing in endoplasmic reticulum
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