Elsevier

Journal of Proteomics

Volume 75, Issue 6, 16 March 2012, Pages 1867-1885
Journal of Proteomics

Comparative proteomic analysis of salt response proteins in seedling roots of two wheat varieties

https://doi.org/10.1016/j.jprot.2011.12.032Get rights and content

Abstract

A comparative proteomic analysis was made of salt response in seedling roots of wheat cultivars Jing-411 (salt tolerant) and Chinese Spring (salt sensitive) subjected to a range of salt stress concentrations (0.5%, 1.5% and 2.5%) for 2 days. One hundred and ninety eight differentially expressed protein spots (DEPs) were located with at least two-fold differences in abundance on 2-DE maps, of which 144 were identified by MALDI-TOF-TOF MS. These proteins were involved primarily in carbon metabolism (31.9%), detoxification and defense (12.5%), chaperones (5.6%) and signal transduction (4.9%). Comparative analysis showed that 41 DEPs were salt responsive with significant expression changes in both varieties under salt stress, and 99 (52 in Jing-411 and 47 in Chinese Spring) were variety specific. Only 15 and 9 DEPs in Jing-411 and Chinese Spring, respectively, were up-regulated in abundance under all three salt concentrations. All dynamics of the DEPs were analyzed across all treatments. Some salt responsive DEPs, such as guanine nucleotide-binding protein subunit beta-like protein, RuBisCO large subunit-binding protein subunit alpha and pathogenesis related protein 10, were up-regulated significantly in Jing-411 under all salt concentrations, whereas they were down-regulated in salinity-stressed Chinese Spring.

Graphical abstract

Highlights

► One highly salt tolerant wheat cultivar and one salt sensitive wheat line were studied. ► Their seedling roots were used for a comparative proteome analysis for salt response. ► 114 DEPs were identified which fall into 4 functional categories.

Introduction

Soil salinity is a prevalent abiotic stress, which seriously impairs crop production on at least 20% of irrigated land worldwide [1]. Salinity stress leads to slow growth, wilting or even death of plants, especially in high salt concentrations. Ion toxicity, nutrient constraints, hyperosmotic stress and oxidative stress caused by salt stress may be the primary causes of severely disrupted protein synthesis and act by interfering with normal enzyme activity [2], [3]. Under salt stress, plants accumulate ion and reactive oxygen species (ROS) that are harmful to plant cells, especially under high salt concentrations [4]. These toxic by-products can decrease enzyme activity or even degrade some proteins. Due to genotypic difference and environmental conditions, plants have adaptive mechanisms to minimize salt injury, and different plants have developed different abilities to survive salt stress. In barley, S-adenosylmethionine (SAM) synthase and peroxidase involved in the detoxification of reactive oxygen species (ROS) were more abundant in salt-tolerant cultivar (cv) Steptoe than in salt-sensitive cv Morex, while proteins involved in iron uptake were expressed at a higher level in the sensitive cv Morex [5]. Salt-tolerant barley cultivars may have greater ability to sequester Na+ into sub-cellular compartments and/or maintain K+ homeostasis during salt stress [6]. Higher levels of vacuolar H+-ATPase might play a pivotal role in salinity tolerance of plant roots.

To date, a number of salt-responsive genes involved in membrane transport, signal transduction, redox reaction and other processes have been identified. Many stress-related genes such as AtNHX1, AVP1, AtSOS1 and AtSOS2 are required for Na+ sequestration and extrusion to maintain intracellular Na+/K+ homeostasis [7], [8], [9]. Such genes also include AgNHX1 and SKC1 in rice [10], and GhNHX1 in tobacco [11]. Some small molecular compatible solutes were synthesized by a series of genes closely related to abiotic stress responses. For example, the BADH gene in Suaeda liaotungensis kitag, which encodes betaine aldehyde dehydrogenase, the key enzyme of glycinebetaine synthesis, was introduced into maize to improve salt tolerance [12], [13], [14]. Sorbitol dehydrogenase encoded by the PmSDH1 gene over-accumulates mannitol to regulate salt stress in transgenic Plantago major plants [7]. Similarly, some transcriptional factors, such as OsNAC5 and JERF3 in rice and TaSTRG in wheat, can regulate stress responses and transgenic plants had improved tolerance to abiotic stresses such as salt, drought and cold [15], [16]. Despite these examples, salinity response remains a very complicated quantitative trait, causing many proteins to undergo removal of signal peptides, RNA splicing and post-translational modifications (PTMs) such as phosphorylation and glycosylation. This leads to poor correlations between transcriptomes and proteomes in different wheat cultivars under abiotic stresses [17], [18].

Recently, proteomic analysis has become one of the best strategies to reveal the dynamics of expression under salt stress. Comparative analysis of root proteomes between two durum wheat varieties with different tolerance levels to NaCl showed that the net synthesis of a 26 kDa polypeptide was significantly changed, being more evident in the more tolerant variety under 200 mM salt stress [19]. Wang et al. [20] identified 49 salt-responsive DEPs between seedling-roots of wheat cultivars Shanrong No. 3 and Jinan 177 under 200 mM salt treatments for 24 h. A wheat V-H+-ATPase E subunit protein was enhanced by salt stress, evidently more so in a wheat salt tolerant cultivar, under 137 mM salt stress [21]. However, proteomic studies on wheat roots at different levels of salt stress are rather limited; the few reports in this area were all focused on a very narrow range of salt concentrations [19], [20], [21]. Thus, comparisons of proteomic dynamics between salt-tolerant and salt-sensitive wheat varieties are yet to be studied, especially under a range of salt concentrations.

Wheat, the second major crop in the world, is a salt-sensitive glycophyte significantly affected by soil salinity. Since the root plays important roles in plant positioning, water absorption, and mineral uptake, it is also considered to be the primary site of salinity perception and the main organ responsible for tolerance to salt stress [2]. A comprehensive survey of the root proteome in response to salinity stress will help in understanding salt tolerance in wheat. Common wheat cv Chinese Spring (CS), which is widely used in experimental studies is sensitive to salt [22] and other stresses such as heat [23]. Chinese cv Jing-411, widely cultivated in the Beijing area in the 1990s, has characteristics of high yield, lodging resistance and abiotic stress tolerance. However, the protein dynamics of salt tolerance have not been investigated. In the present work, we undertook a comparative proteomic analysis of roots of wheat cv Jing-411 and Chinese Spring after exposure to a gradient of salinity conditions.

Section snippets

Plant materials and salt treatment

The experiments were carried out on common wheat varieties (Triticum aestivum L., 2n = 6x = 42, AABBDD), Jing-411 and Chinese Spring. Seeds were germinated on wet filter paper in the dark at room temperature. Uniformly pregerminated seeds were grown in 16 h light and 8 h dark at 23 °C–25 °C. Two-leaf seedlings were transferred to Hoagland's solution containing 5 mM KNO3, 2 mM MgSO4, 1 mM KH2PO4, 5 mM Ca(NO3)2, 50 μM FeNa2(EDTA)2, 50 μM H3BO3, 10 μM MnC12, 0.8 μM ZnSO4, 0.4 μM CuSO4, and 0.02 μM (NH4)6MoO24. The

Morphological and physiological changes under salt stress

In order to study short-term responses to salt stress, three-week-old wheat seedlings of the two wheat cultivars were treated for 2 days under a gradient of salt concentrations. CS wilted more than Jing-411 and exhibited obvious chlorosis, especially at the highest salt concentration. After salt treatment, the relative water content (RWC) and chlorophyll content in leaves were decreased more in CS than in Jing-411 (Fig. 1). The sodium contents in the roots differed significantly between CS and

Discussion

Our results demonstrated that, after treatment for 2 days under a gradient of salt concentrations, CS wilted more to a greater extent than Jing-411 and exhibited obvious chlorosis. Salt stress inhibits plant growth for two main reasons; it reduces the ability of the plant to take up water (osmotic stress) and it accumulates to excessive levels in the tissues resulting in cellular injury (ionic stress) [25], [26]. Under high salt concentrations, the regulatory functions of the plant appear to be

Summary

Salt tolerance is a complex phenomenon in most plant species, and involves numerous mechanisms, at the cellular, tissue, organ, and whole plant levels [62]. In the current study, a significant number of salt tolerance-related proteins were identified with various functions, including signal transduction related proteins, carbon, amino acid and nitrogen metabolism proteins, and detoxification and defense-associated proteins. These proteins should be useful in revealing insights into salt

Acknowledgments

We are grateful to Professor Robert McIntosh from University of Sydney for constructive suggestions in reviewing the manuscript. This research was financially supported by grants from the National Natural Science Foundation of China (30830072), the Chinese Ministry of Science and Technology (2009CB118300) and Key Project of National Plant Transgenic Genes of China (2008ZX08002-004, 2009ZX08002-017B).

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    These authors contributed equally to this work.

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