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Mol Cell Proteomics. 2018 Oct;17(10):2068-2080. doi: 10.1074/mcp.TIR118.000702. Epub 2018 Jul 13.

Universal Plant Phosphoproteomics Workflow and Its Application to Tomato Signaling in Response to Cold Stress.

Author information

1
From the ‡Department of Biochemistry, Purdue University, West Lafayette, IN 47907.
2
§Department of Horticulture and Landscape, Purdue University, West Lafayette, IN 47907.
3
¶Shanghai Center for Plant Stress Biology and Center of Excellence in Molecular Plant Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
4
‖Institute of Plant Stress Biology, State Key Laboratory of Cotton Biology, Department of Biology, Henan University, Kaifeng 475001, China.
5
**Department of Chemistry, Purdue University, West Lafayette, IN 47907.
6
From the ‡Department of Biochemistry, Purdue University, West Lafayette, IN 47907; watao@purdue.edu.

Abstract

Phosphorylation-mediated signaling transduction plays a crucial role in the regulation of plant defense mechanisms against environmental stresses. To address the high complexity and dynamic range of plant proteomes and phosphoproteomes, we present a universal sample preparation procedure that facilitates plant phosphoproteomic profiling. This advanced workflow significantly improves phosphopeptide identifications, enabling deep insight into plant phosphoproteomes. We then applied the workflow to study the phosphorylation events involved in tomato cold tolerance mechanisms. Phosphoproteomic changes of two tomato species (N135 Green Gage and Atacames) with distinct cold tolerance phenotypes were profiled under cold stress. In total, we identified more than 30,000 unique phosphopeptides from tomato leaves, representing about 5500 phosphoproteins, thereby creating the largest tomato phosphoproteomic resource to date. The data, along with the validation through in vitro kinase reactions, allowed us to identify kinases involved in cold tolerant signaling and discover distinctive kinase-substrate events in two tomato species in response to a cold environment. The activation of SnRK2s and their direct substrates may assist N135 Green Gage tomatoes in surviving long-term cold stress. Taken together, the streamlined approach and the resulting deep phosphoproteomic analyses revealed a global view of tomato cold-induced signaling mechanisms.

KEYWORDS:

Cold Stress; Mass Spectrometry; Phosphoproteome; Phosphorylation; Signal Transduction; Stress response; Tomato Phosphoproteome

PMID:
30006488
PMCID:
PMC6166681
[Available on 2019-10-01]
DOI:
10.1074/mcp.TIR118.000702

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