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作物学报 ›› 2020, Vol. 46 ›› Issue (8): 1275-1282.doi: 10.3724/SP.J.1006.2020.91068

• 研究简报 • 上一篇    下一篇

两个不同籽粒硬度小麦的比较蛋白组学分析

刘培勋1,马小飞2,万洪深1,郑建敏1,罗江陶1,蒲宗君1,*()   

  1. 1四川省农业科学院作物研究所/农业农村部西南地区小麦生物学与遗传育种重点实验室, 四川成都 610066
    2山西省农业科学院小麦研究所, 山西临汾 041000
  • 收稿日期:2019-11-23 接受日期:2020-03-30 出版日期:2020-08-12 网络出版日期:2020-05-17
  • 通讯作者: 蒲宗君
  • 作者简介:E-mail: littlefarmer@163.com; Tel: 028-84504231
  • 基金资助:
    国家自然科学基金项目(31671683);国家自然科学基金项目(31401383);四川省财政创新能力提升工程(2016ZYPZ-016);四川省财政创新能力提升工程(2019QNJJ-007);四川省财政创新能力提升工程(2019QYXK034);四川省科技计划项目(2017JY0077)

Comparative proteomic analysis of two wheat genotypes with contrasting grain softness index

LIU Pei-Xun1,MA Xiao-Fei2,WAN Hong-Shen1,ZHENG Jian-Min1,LUO Jiang-Tao1,PU Zong-Jun1,*()   

  1. 1Crop Research Institute, Sichuan Academy of Agricultural Sciences/Key Laboratory of Wheat Biology and Genetic Improvement on Southwestern China, Ministry of Agriculture and Rural Areas, Chengdu 610066, Sichuan, China
    2Wheat Research Institute, Shanxi Academy of Agricultural Sciences, Linfen 041000, Shanxi, China
  • Received:2019-11-23 Accepted:2020-03-30 Published:2020-08-12 Published online:2020-05-17
  • Contact: Zong-Jun PU
  • Supported by:
    National Natural Science Foundation of China(31671683);National Natural Science Foundation of China(31401383);Financial Innovation Capacity Improvement Project of Sichuan Province(2016ZYPZ-016);Financial Innovation Capacity Improvement Project of Sichuan Province(2019QNJJ-007);Financial Innovation Capacity Improvement Project of Sichuan Province(2019QYXK034);Science and Technology Planning Project of Sichuan Province(2017JY0077)

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摘要：

小麦是全球种植面积最大粮食作物, 为全球45亿人提供日常蛋白和能量摄入的20%。弄清小麦籽粒硬度遗传基础, 对于改良小麦品质具有重大意义。为探讨不同硬度小麦种子的分子基础, 本实验选用西南麦区2个硬度差异极显著的小麦品种川麦66和蜀麦969, 从蛋白水平上分析其种子蛋白差异表达情况, 利用TMT定量蛋白质组学技术(tandem mass tags)结合生物信息学分析, 分析差异表达的蛋白及其功能和通路等富集情况。结果表明, 鉴定并定量了有效蛋白6020个, 其中显著差异表达蛋白113个, 在软质麦川麦66中上调表达的69个, 下调表达的44个。差异蛋白GO富集分析共富集到65个GO条目, 达到极显著富集水平的包括生物过程的1个条目、细胞组成的1个条目和分子功能的6个条目。推测营养库活性类蛋白、酶抑制剂活性类蛋白和谷胱甘肽代谢途径类蛋白可能参与小麦籽粒硬度形成。籽粒硬度相关蛋白可能主要分布于细胞胞外区, 具有防御作用。从系统发育分析推测, puroindolines蛋白及其同源蛋白, 可能既作为小麦籽粒贮藏蛋白, 同时还能作为酶抑制剂调控籽粒发育。本研究为进一步探索小麦籽粒硬度遗传机制提供了参考。

关键词: 小麦, 籽粒硬度, TMT, 蛋白质组

Abstract:

Wheat is the crop most widely grown in the world and provides the daily protein and 20% food calories for 4.5 billion people. It is crucial to understand the genetic basis of grain hardness for improving wheat quality. In order to explore the molecular basis of the formation of wheat grain hardness, two wheat cultivars Chuanmai 66 and Shumai 969 with significant hardness difference in southwest wheat region were selected to analyze the proteins differential expression by TMT quantitative proteomics (tandem mass tags) and bioinformatic methods of function and pathway enrichment analysis. A total of 6020 effective proteins were identified and quantified, including 113 differentially expressed proteins (DEPs), of which 69 were up-regulated and 44 were down-regulated in soft wheat Chuanmai 66. These DEPs were enriched into 65 GO terms, including a biological process term, a cellular component term and six molecular function terms at extremely significant level. Based on the enrichment analysis, we suggested that nutrient reservoir activity proteins, enzyme inhibitor proteins and glutathione metabolism proteins might participate in the formation of wheat grain hardness, and grain hardness related proteins might mainly distribute in the extracellular region of cells and had defensive function. According to the phylogenetic analysis, it was inferred that puroindolines and its homologous proteins might be as not only wheat grain storage proteins, but also enzyme inhibitors regulating grain development. This study provides a basis for further exploring the genetic mechanism of wheat grain hardness.

Key words: Triticum aestivum L., grain hardness, TMT, protemics

图1

川麦66和蜀麦969籽粒主要品质指标比较"

图2

蛋白质功能注释结果"

图3

差异表达蛋白火山图"

图4

GO富集柱状图 A属于生物过程(BP), B属于细胞组分(CC), C、D、E、F、G、H、I、J属于分子功能(MF)。A: 防御反应; B: 胞外区; C: 营养库活性; D: 酶抑制剂活性; E: 酶调节活性; F: 丝氨酸型内肽酶抑制剂活性; G: 肽链内切酶抑制剂活性; H: 天冬氨酸型内肽酶活性; I: 电子载体活性; J: 几丁质结合。"

表1

营养库活性类相关差异表达蛋白(川麦66 vs. 蜀麦969)"

蛋白名称
Protein ID		 描述
Description		 差异倍数
Fold change		 上调/下调
Up/down
B2Y2Q6 B2Y2Q6_WHEAT LMW-B2 1.5425 Up
B2BZC7 B2BZC7_WHEAT LMW-m glutenin subunit 0154A5-M 1.8737 Up
A0A173DQZ1 A0A173DQZ1_WHEAT type-b avenin-like protein 3.8163 Up
Q8H0J5 Q8H0J5_WHEAT low molecular weight glutenin subunit (fragment) 5.0056 Up
TraesCS7A01G035300.1 Gliadin-like avenin 2.9853 Up
Q6WZC3 Q6WZC3_WHEAT low molecular weight glutenin subunit 2.1622 Up
TraesCS4A01G453400.1 Gamma-gliadin 7.2606 Up
A0A286QTK1 A0A286QTK1_WHEAT avenin-like protein A2 0.5984 Down
F8SGL3 F8SGL3_WHEAT low-molecular-weight glutenin subunit 0.5493 Down
Q0QBR3 Q0QBR3_WHEAT LMW-glutenin P3-5 0.3190 Down
B2Y2R3 B2Y2R3_WHEAT low molecular weight glutenin subunit 0.5725 Down
B6UKN9 B6UKN9_WHEAT gamma-gliadin 0.6340 Down
TraesCS2A01G211800.1 Germin-like protein 1-1 0.5875 Down
A0A2P1H6A2 A0A2P1H6A2_WHEAT alpha-gliadin 0.5908 Down
A0A0K2QJY6 A0A0K2QJY6_WHEAT alpha/beta-gliadin 0.6081 Down
I0IT65 I0IT65_WHEAT alpha-gliadin 0.6241 Down

表2

酶抑制剂类相关差异表达蛋白(川麦66 vs. 蜀麦969)"

蛋白名称
Protein ID		 描述
Description		 差异倍数
Fold change		 上调/下调
Up/down
Q5UHH6 Q5UHH6_WHEAT 0.19 dimeric alpha-amylase inhibitor (fragment) 1.6788 Up
TraesCS5D01G004300.1 Puroindoline-b, protease inhibitor/seed storage/LTP family 2.8227 Up
P81713 IBB3_WHEAT Bowman-Birk type trypsin inhibitor 1.5092 Up
TraesCS4A01G460900.1 Invertase inhibitor, plant invertase/pectin methylesterase inhibitor 1.5169 Up
TraesCS1D01G028500.1 Chymotrypsin inhibitor 2.3465 Up
TraesCS5D01G561800.1 Invertase inhibitor, plant invertase/pectin methylesterase inhibitor 2.0582 Up
A0A080YTU1 A0A080YTU1_WHEAT uncharacterized protein 1.5779 Up
A0A2X0S1F0 A0A2X0S1F0_WHEAT peptidase A1 domain-containing protein 1.9149 Up
TraesCS4D01G205800.1 ADP-ribosylation factor GTPase-activating protein 1.6650 Up
TraesCS7A01G502500.1 Eukaryotic aspartyl protease family protein 2.3542 Up
TraesCS3D01G467500.1 Eukaryotic aspartyl protease family protein 0.3700 Down
TraesCS4D01G250000.1 Dimeric alpha-amylase inhibitor 0.3659 Down
TraesCS1D01G265900.1 Wound-induced protease inhibitor 0.5474 Down
TraesCS3D01G025700.1 Trypsin inhibitor 0.1974 Down
TraesCS6B01G407700.1 Aspartic proteinase nepenthesin-1 0.6473 Down

表3

谷胱甘肽代谢途径相关差异表达蛋白(川麦66 vs. 蜀麦969)"

蛋白名称
Protein ID		 描述
Description		 差异倍数
Fold change		 上调/下调
Up/down
TraesCS3D01G491400.1 6-phosphogluconate dehydrogenase 2.5968 Up
TraesCS2B01G096200.1 Ascorbate peroxidase 1.5695 Up
TraesCS3A01G488200.1 Glutathione S-transferase 1.5215 Up
TraesCS3B01G536100.1 Glutathione S-transferase 1.9499 Up
TraesCS3D01G445400.1 Glutathione S-transferase 0.6058 Down
W5D4D9 W5D4D9_WHEAT uncharacterized protein 0.5675 Down

图5

软质麦中上调表达蛋白与puroindolines蛋白共同构建的系统发育树 带实心圆形的为puroindolines蛋白; 带空心正方形的为营养库活性类别; 带空心三角形的为酶抑制剂活性类别; 带空心圆形的为谷胱甘肽代谢途径蛋白。"

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