Quantitative redox proteomics revealed molecular mechanisms of salt tolerance in the roots of sugar beet monomeric addition line M14

Abstract

Background: Salt stress is often associated with excessive production of reactive oxygen species (ROS). Oxidative stress caused by the accumulation of ROS is a major factor that negatively affects crop growth and yield. Root is the primary organ that senses and transmits the salt stress signal to the whole plant. How oxidative stress affect redox sensitive proteins in the roots is not known.

Results: In this study, the redox proteome of sugar beet M14 roots under salt stress was investigated. Using iTRAQ reporters, we determined that salt stress caused significant changes in the abundance of many proteins (2305 at 20 min salt stress and 2663 at 10 min salt stress). Using iodoTMT reporters, a total of 95 redox proteins were determined to be responsive to salt stress after normalizing again total protein level changes. Notably, most of the differential redox proteins were involved in metabolism, ROS homeostasis, and stress and defense, while a small number play a role in transport, biosynthesis, signal transduction, transcription and photosynthesis. Transcription levels of 14 genes encoding the identified redox proteins were analyzed using qRT-PCR. All the genes were induced by salt stress at the transcriptional level.

Conclusions: Based on the redox proteomics results, we construct a map of the regulatory network of M14 root redox proteins in response to salt stress. This study further refines the molecular mechanism of salt resistance at the level of protein redox regulation.

Keywords: Molecular mechanisms; Redox proteomics; Salt stress; Sugar beet M14; iodoTMTRAQ.

Fig. 1

Temporal changes in cysteine free sulfhydryl, AsA, and GSH contents in BvM14 roots under salt stress. A Cysteine free sulfhydryl content under 200 mM and 400 mM NaCl stress. B ASA content under 200 mM and 400 mM NaCl stress. C GSH content under 200 mM and 400 mM NaCl stress. These values are the means of three biological replicates from different samples with standard errors. *p < 0.05; **p < 0.01

Fig. 2

Visualization of redox protein profile data from BvM14 roots under salt stress. A iTRAQ-labeled total protein and iodoTMT-labeled redox protein of BvM14 under 200 mM and 400 mM NaCl stress. B Significant changes in protein redox levels in BvM14 roots under salt stress. C Comparison of the number of differential redox proteins identified under 200 mM NaCl and 400 mM NaCl treatments

Fig. 3

Functional classification and subcellular localization of the differential redox proteins. A Functional classification of the differential redox proteins. B Subcellular localization prediction of the differential redox proteins. C Number of redox proteins in each function

Fig. 4

Comparative analysis of differential redox proteins in sugar beet M14 roots and leaves under salt stress. A Comparative analysis of redox protein functions under salt stress in roots and leaves. B Comparison of protein redox levels under salt stress in roots and leaves of the M14. EG45 EG45-like domain containing protein, RD19A cysteine protease RD19A, NADH NADH dehydrogenase [ubiquinone] 1 alpha, VSR vacuolar-sorting receptor, GSAM glutamate-1-semialdehyde 2,1-aminomutase, Fd ferredoxin, root R-B1, Pfn profilin, POD peroxidase, Hsp heat shock cognate protein

Fig. 5

Real-Time PCR assays of genes encoding differential redox proteins and differential proteins in different pathways. A RealTime PCR assays of genes encoding redox proteins common to roots and leaves under salt stress, B RealTime PCR assays of genes encoding redox proteins specific to 200 mM or 400 mM salt stress condition, and C Real-Time PCR assays of genes encoding redox proteins common to 200 mM and 400 mM salt stress. The x-axis is the salt concentration. y-Axis is the relative expression of each gene (2^−ΔΔCT). Please refer to Table 1 for abbreviations

Fig. 6

The metabolic networks of the redox protein in sugar beet M14 roots under salt stress. Under 200 mM NaCl treatment, the reduced protein is orange colors and the oxidized protein is green colors. Under 400 mM NaCl treatment, the reduced protein is red colors and the oxidized protein is blue colors. Please refer to Additional file 7: Table S6 for abbreviations. The black underline represents redox proteins common to both leaves and roots

Fig. 7

Alignment of amino acid sequence of different expression of peroxidase in salt stress response. Black boxes indicate conserved Cys sites and red boxes indicate Cys sites that undergo redox modifications