GENE EXPRESSION CHANGES DURING SAR
There is ample evidence indicating that SAR is conferred by expression of a collection of genes. The phenotype of dth9 is a good example, showing that SAR can be blocked without affecting the induction of PR-1 and PR-2. The sequencing of the Arabidopsisgenome has allowed global analyses of gene expression changes during SAR to be conducted using DNA microarray technology. Researchers surveyed 25–30% of Arabidopsis genes under 14 SAR-inducing or repressing conditions, identifying 413 ESTs (about 300 genes) that show differential expression during SAR (65). Cluster analysis was used to identify a group of 31 genes with a similar transcription pattern to that of PR-1. The genes in this PR-1 regulon are strongly induced in systemic tissue during SAR and this induction is NPR1-dependent. They are also induced by infection with virulent P. parasitica, suggesting that activation of SAR-related genes in local tissue can limit infection by compatible pathogens. This is consistent with the enhanced susceptibility to virulent pathogens observed in npr1 mutants. Interestingly, only 17 of 26 PR-1 regulon genes have an as-1 element in their promoter, whereas W-boxes were overrepresented, occurring an average of 4.3 times in every promoter. This suggests that WRKY factors rather than TGA factors are important for coregulation of PR-1 regulon genes. The LS4 W-box in the PR-1 promoter acts as a strong negative cis-element, leading Maleck et al. to propose that WRKY factors repress the expression of PR-1 regulon genes. Upon activation of SAR, NPR1-dependent derepression would occur, possibly through the inactivation of SNI1.
Promoter analysis was also performed in another study on 1058 genes that were induced by pathogen infection, SA, MeJA, or ozone. This revealed that as-1 elements, W-boxes, abscisic acid response elements, and G-boxes were overrepresented across all treatments, whereas the Myc motif was overrepresented only in the SA-induced genes. This suggests a role for these cis-elements in stress responses but does not identify which are important during SAR.
A potential difficulty in interpreting microarray data is the lack of hierarchical information on the transcriptional events that occur during SAR. The SAR-induced genes include effector genes that confer resistance as well as regulatory genes such as transcription factors. To overcome this problem, a strategy was devised to focus on only a single transcriptional level (D. Wang & X. Dong, unpublished data). Fusion of NPR1 to the hormone-binding domain of the rat glucocorticoid receptor (GR) placed its nuclear localization under the control of the steroid hormone dexamethasone. Treating an NPR1-GR transgenic line (in npr1 background) with SA will activate expression of SA-mediated NPR1-independent genes. Addition of dexamethasone after SA treatment will allow NPR1-GR to move to the nucleus and induce NPR1-mediated genes. To focus on the direct transcriptional targets of NPR1, cycloheximide can be added to inhibit de novo protein synthesis and secondary transcriptional events.