Comparative Analysis Of Salt Stress-responsive Glucosinolate Profile And Proteome In Arabidopsis Thaliana And Thellungiella Halophila

Here we report glucosinolate profile and comparative proteomics of Arabidopsis thaliana, a glycophyte, and its close relative Thellungiella halophila, a halophyte, under different salt stress conditions. Soluble proteins from control and NaCl treated samples were extracted and separated by two-dimensional gel electrophoresis. As a complementary approach, isobaric tag for relative and absolute quantification (iTRAQ) LC-MS was used to identify crude microsomal proteins. Collectively, this work represents the most extensive proteomic description of salinity responses of A. thaliana and T. halophila. The studies may provide the basis for the future research of the effect of salt stress on glucosinolate metabolism and salt tolerance molecular mechanism in glycophytes and halophytes.First, seven glucosinolates in T. halophila were unambiguously identified by high performance liquid chromatography and mass spectrometry (HPLC-MS) for the first time, including aliphatic glucosinolates Allyl,3MSOP,10MSD and 3MTP, indole glucosinolates I3M and 4MO13M, aromatic glucosinolate 2PE. The composition of glucosinolates varied considerably among different tissues, with seven compounds identified in flowers, siliques, cotyledons and roots, six in stems, cauline leaves and petioles (I3M was not detected), five in rosette leaves (2PE and I3M were not detected), only four in seeds and all four glucosinolates belong to aliphatic (including Allyl,3MSOP,10MSD and 3MTP). The total contents of glucosinolates in different organs at various developmental stages also displayed significant variationsSecond, the composition of glucosinolates was not varied in the rosette leaves of four-week-old A. thaliana and 6-week-old T. halophila, which were irrigated with 50 mM and 150 mM NaCl. The contents of total, aromatic and indole glucosinolates were not varied significantly in A. thaliana after treatment. However, the contents of total, aromatic and indole glucosinolates decreased after 50 mM NaCl treated and increased higher than control after 150 mM NaCl treated. Aliphatic glucosinolates 3MSOP,5MSOP and indole glucosinolates 4OHI3M and 4MOI3M in A. thaliana decreased with increasing NaCl concentrations, while aliphatic glucosinolates 6MSOH, indole glucosinolates I3M in A. thaliana and aliphatic glucosinolates 3MSOP in T. halophila increased with increasing NaCl concentrations. The content of 8MSOO and 1MOI3M in A. thaliana,3MTP, Allyl,10MSD and 4MOI3M in T. halophila displayed a similar pattern of variation.Third, soluble proteins from control and NaCl treated samples were separated by two-dimensional gel electrophoresis. A total of 88 protein spots from A. thaliana gels and 37 protein spots from T. halophila gels showed significant changes. Out of these spots, a total of 79 and 32 proteins, which were significantly regulated by salt stress, were identified by mass spectrometry in A. thaliana and T. halophila, respectively. Most of the changes were observed in A. thaliana samples treated with 150 mM NaCl. In contrast, most proteins were found to exhibit significantly changes in 50 mM NaCl treated T. halophila samples.Fourth, isobaric tag for relative and absolute quantification (iTRAQ) LC-MS was used to identify crude microsomal proteins. There were 31 differentially expressed (21 increased and 10 decreased) in A. thaliana. In T. halophila, there were 32 differentially expressed proteins (11 increased and 21 decreased). Most of the differentially expressed proteins are membrane proteins or membrane associated proteins in chloroplasts, mitochondria and plasma membraneFifth, the non-redundant identified proteins were grouped according to functional categories. In T. halophila the proteins were grouped into 11 categories, the largest group is the protein synthesis (22%). In A. thaliana, the salt-responsive proteins were classified into 13 categories, metabolic proteins (22%) formed the largest group. This finding suggests that although proteins involved in plant metabolism play an important role in A. thaliana salt tolerance, the protein synthesis-related proteins play an important role in T. halophila salt tolerance.