| Salt stress is one of the dominant abiotic stresses that influence plant growth and decrease the harvestable yield of all crop species. Roots are the primary site of salinity perception. To better understand salt stress responses in Arabidopsis roots, both the transcriptome and proteome changes in Arabidopsis roots treated by 150 mM NaC1 for short and long terms were monitored using a full-genome oligonucleotide microarray and two-dimensional gel electrophoresis (2-DGE) based technologies, respectively. Statistical analysis of microarray results revealed that 5, 280 (22%) genes were salt-regulated with 2, 367 genes up-regulated and 2, 913 genes down-regulated by a cutoff of 2-fold. Many novel salt-responsive genes including transporters and transcription factors (TFs) were identified. A comparative proteomic analysis of salt-stressed Arabidopsis roots using 2-DGE and liquid-chromatography-tandem mass spectrometry (LC-MS/MS) allowed identification of 86 differentially expressed proteins, including many previously characterized stress-responsive proteins and proteins implicated in reactive oxygen species (ROS) scavenging, signal transduction, translation, cell wall biosynthesis as well as energy, amino acid, and hormone metabolisms. These data confirmed the modest correlation between mRNA and protein levels. Three novel salt-inducible transcription factor genes, bHLH92, WRKY25 and WRKY33 were chosen for further characterization through a combination of reverse genetic strategies including knock-out and overexpression. Quantitative RT-PCR demonstrated that the three TFs were responsive to multiple abiotic stresses. Promoter-GUS fusion assays revealed the spatial and developmental expression patterns and demonstrated that promoters are inducible by some abiotic stresses. Knock-out mutants for the three TFs were identified, and stress tolerance assays showed that null mutants displayed minor differences from wild-type plants under some specific abiotic stresses, suggesting a functional redundancy exists between members of the same gene families. Constitutive overexpression of the three TFs in Arabidopsis enhanced its tolerance to salt and/or osmotic stress. Finally microarrays were employed to identify the target genes of the three TFs under saline condition, and many target genes were found to be differentially expressed between mutants and wild-type. This study not only provides a more comprehensive description of salt-responsive genes and proteins, but also contributes to elucidating the roles of bHLH and WRKY TFs in abiotic stress. |
