title: Mechanisms of Soybean Roots' Tolerances to Salinity Revealed by Proteomic and Phosphoproteomic Comparisons Between Two Cultivars.
abstract: Understanding molecular mechanisms underlying plant salinity tolerance provides valuable knowledgebase for effective crop improvement through genetic engineering. Current proteomic technologies, which support reliable and high-throughput analyses, have been broadly used for exploring sophisticated molecular networks in plants. In the current study, we compared phosphoproteomic and proteomic changes in roots of different soybean seedlings of a salt-tolerant cultivar (Wenfeng07) and a salt-sensitive cultivar (Union85140) induced by salt stress. The root samples of Wenfeng07 and Union85140 at three-trifoliate stage were collected at 0 h, 0.5 h, 1 h, 4 h, 12 h, 24 h, and 48 h after been treated with 150 mm NaCl. LC-MS/MS based phosphoproteomic analysis of these samples identified a total of 2692 phosphoproteins and 5509 phosphorylation sites. Of these, 2344 phosphoproteins containing 3744 phosphorylation sites were quantitatively analyzed. Our results showed that 1163 phosphorylation sites were differentially phosphorylated in the two compared cultivars. Among them, 10 MYB/MYB transcription factor like proteins were identified with fluctuating phosphorylation modifications at different time points, indicating that their crucial roles in regulating flavonol accumulation might be mediated by phosphorylated modifications. In addition, the protein expression profiles of these two cultivars were compared using LC MS/MS based shotgun proteomic analysis, and expression pattern of all the 89 differentially expressed proteins were independently confirmed by qRT-PCR. Interestingly, the enzymes involved in chalcone metabolic pathway exhibited positive correlations with salt tolerance. We confirmed the functional relevance of chalcone synthase, chalcone isomerase, and cytochrome P450 monooxygenase genes using soybean composites and Arabidopsis thaliana mutants, and found that their salt tolerance were positively regulated by chalcone synthase, but was negatively regulated by chalcone isomerase and cytochrome P450 monooxygenase. A novel salt tolerance pathway involving chalcone metabolism, mostly mediated by phosphorylated MYB transcription factors, was proposed based on our findings. (The mass spectrometry raw data are available via ProteomeXchange with identifier PXD002856).
authors.affiliation: From the ‡College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, Zhejiang, 310036, PR China; pierzaixian001@aliyun.com liqundu@hznu.edu.cn smngai@cuhk.edu.hk.
authors.forename: Erxu
authors.initials: E
authors.lastname: Pi
authors.affiliation: From the ‡College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, Zhejiang, 310036, PR China;
authors.forename: Liqun
authors.initials: L
authors.lastname: Qu
authors.affiliation: §Shanghai Applied Protein Technology Co. Ltd, Shanghai, 200233, PR China;
authors.forename: Jianwen
authors.initials: J
authors.lastname: Hu
authors.affiliation: From the ‡College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, Zhejiang, 310036, PR China;
authors.forename: Yingying
authors.initials: Y
authors.lastname: Huang
authors.affiliation: ¶The National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI), Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing, P.R. China;
authors.forename: Lijuan
authors.initials: L
authors.lastname: Qiu
authors.affiliation: ‖College of Life Science, Zhejiang Sci-Tech University, Hangzhou, 310018, PR China;
authors.forename: Hongfei
authors.initials: H
authors.lastname: Lu
authors.affiliation: **College of Biology and Food Engineering, Changshu Institute of Technology, Changshu 215500, PR China;
authors.forename: Bo
authors.initials: B
authors.lastname: Jiang
authors.affiliation: From the ‡College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, Zhejiang, 310036, PR China;
authors.forename: Cong
authors.initials: C
authors.lastname: Liu
authors.affiliation: From the ‡College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, Zhejiang, 310036, PR China;
authors.forename: Tingting
authors.initials: T
authors.lastname: Peng
authors.affiliation: From the ‡College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, Zhejiang, 310036, PR China;
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: From the ‡College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, Zhejiang, 310036, PR China;
authors.forename: Huizhong
authors.initials: H
authors.lastname: Wang
authors.affiliation: ‡‡Centre for Soybean Research of Partner State Key Laboratory of Agrobiotechnology and School of Life Sciences, The Chinese University of Hong Kong, Hong Kong.
authors.forename: Sau-Na
authors.initials: SN
authors.lastname: Tsai
authors.affiliation: ‡‡Centre for Soybean Research of Partner State Key Laboratory of Agrobiotechnology and School of Life Sciences, The Chinese University of Hong Kong, Hong Kong pierzaixian001@aliyun.com liqundu@hznu.edu.cn smngai@cuhk.edu.hk.
authors.forename: Saiming
authors.initials: S
authors.lastname: Ngai
authors.affiliation: From the ‡College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, Zhejiang, 310036, PR China; pierzaixian001@aliyun.com liqundu@hznu.edu.cn smngai@cuhk.edu.hk.
authors.forename: Liqun
authors.initials: L
authors.lastname: Du
mesh_terms: D000217:Acyltransferases / Q000235:genetics / Q000378:metabolism; D002853:Chromatography, Liquid; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D020869:Gene Expression Profiling / Q000379:methods; D019753:Intramolecular Lyases / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D018517:Plant Roots / Q000235:genetics / Q000378:metabolism*; D020543:Proteome / Q000235:genetics / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055049:Salt Tolerance / Q000235:genetics; D013025:Glycine max / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D013045:Species Specificity; D053719:Tandem Mass Spectrometry; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D010750:Phosphoproteins; D010940:Plant Proteins; D020543:Proteome; D014157:Transcription Factors; D003577:Cytochrome P-450 Enzyme System; D000217:Acyltransferases; C020141:flavanone synthetase; D019753:Intramolecular Lyases; C019865:chalcone isomerase