Proteomic analysis of salt-responsive ubiquitin-related proteins in rice roots

Authors


C.-S. Wang, Department of Agronomy, National Chung-Hsing University, 250 Kuo-Kuang Road, Taichung 40227, Taiwan.

E-mail: wangchansen@nchu.edu.tw

C.-C. Lai, Institute of Molecular Biology, National Chung-Hsing University, 250 Kuo-Kuang Road, Taichung 40227, Taiwan.

E-mail: lailai@dragon.nchu.edu.tw

Abstract

RATIONALE

Ubiquitination of proteins plays an important role in regulating a myriad of physiological functions in plants such as xylogenesis, senescence, cell cycle control, and stress response. However, only a limited number of proteins in plants have been identified as being ubiquitinated in response to salt stress. The relationships between ubiquitination and salt-stress responses in plants are not clear.

METHODS

Rice (Oryza sativa) seedlings from the same genetic background with various salt tolerances exposed to salt stress were studied. The proteins of roots were extracted then analyzed using western blotting against ubiquitin. Differentially expressed ubiquitinated proteins were identified by nanospray liquid chromatography/tandem mass spectrometry (nano-LC/MS/MS) and quantified by label-free methods based on the Exponentially Modified Protein Abundance Index (emPAI) and on the peak areas of XIC spectra derived from ubiquitinated peptides. In addition, we performed a gel-based shotgun proteomic analysis to detect the ubiquitinated proteome that may be involved in response to salt stress.

RESULTS

The expressions of ubiquitination on pyruvate phosphate dikinase 1, heat shock protein 81–1, probable aldehyde oxidase 3, plasma membrane ATPase, cellulose synthase A catalytic subunit 4 [UDP-forming] and cyclin-C1-1 were identified and compared before and after salt treatment. The functions of those ubiquitinated proteins were further discussed for defence against salt stress. In addition, a large number of ubiquitinated proteins were successfully identified as well in this study.

CONCLUSIONS

The ubiquitination of proteins affected the protective mechanisms in rice seedlings to resist the salt stress during the initial phase. The findings in the present study also demonstrate that the regulated mechanisms through protein ubiquitination are important for rice seedlings against salt stress. Copyright © 2012 John Wiley & Sons, Ltd.

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