SummarySalicylic acid (SA) is a plant signaling molecule that mediates the induction of defense responses upon attack by a variety of pathogens. Moreover, it antagonizes gene induction by the stress signaling molecule jasmonic acid (JA). Several SA-responsive genes are regulated by basic/leucine zipper-type transcription factors of the TGA family. TGA factors interact with NPR1, a central regulator of many SA-induced defense responses including SA/JA antagonism. In order to identify further regulatory proteins of SA-dependent signaling pathways, a yeast protein interaction screen with tobacco TGA2.2 as bait and an Arabidopsis thaliana cDNA prey library was performed and led to the identification of a member of the glutaredoxin family (GRX480, encoded by At1g28480). Glutaredoxins are candidates for mediating redox regulation of proteins because of their capacity to catalyze disulfide transitions. This agrees with previous findings that the redox state of both TGA1 and NPR1 changes under inducing conditions. Transgenic Arabidopsis plants ectopically expressing GRX480 show near wild-type expression of standard marker genes for SA-and xenobiotic-inducible responses. In contrast, transcription of the JA-dependent defensin gene PDF1.2 was antagonized by transgenic GRX480. This, together with the observation that GRX480 transcription is SA-inducible and requires NPR1, suggests a role of GRX480 in SA/JA cross-talk. Suppression of PDF1.2 by GRX480 depends on the presence of TGA factors, indicating that the GRX480/TGA interaction is effective in planta.

Tri-methylated H3 lysine 4 (H3K4me3) is associated with transcriptionally active genes, but its function in the transcription process is still unclear. Point mutations in the catalytic domain of ATX1 (ARABIDOPSIS TRITHORAX1), a H3K4 methyltransferase, and RNAi knockdowns of subunits of the AtCOMPASS–like (Arabidopsis Complex Proteins Associated with Set) were used to address this question. We demonstrate that both ATX1 and AtCOMPASS–like are required for high level accumulation of TBP (TATA-binding protein) and Pol II at promoters and that this requirement is independent of the catalytic histone modifying activity. However, the catalytic function is critically required for transcription as H3K4me3 levels determine the efficiency of transcription elongation. The roles of H3K4me3, ATX1, and AtCOMPASS–like may be of a general relevance for transcription of Trithorax-activated eukaryotic genes.

Tightly balanced antagonism between the Polycomb group (PcG) and the Trithorax group (TrxG) complexes maintain Hox expression patterns in Drosophila and murine model systems. Factors belonging to the PcG/TrxG complexes control various processes in plants as well but whether they participate in mechanisms that antagonize, balance or maintain each other's effects at a particular gene locus is unknown. CURLY LEAF (CLF), an Arabidopsis homolog of enhancer of zeste (EZ) and the ARABIDOPSIS HOMOLOG OF TRITHORAX (ATX1) control the expression of the flower homeotic gene AGAMOUS (AG). Disrupted ATX1 or CLF function results in misexpression of AG, recognizable phenotypes and loss of H3K4me3 or H3K27me3 histone H3-tail marks, respectively. A novel idea suggested by our results here, is that PcG and TrxG complexes function as a specific pair generating bivalent chromatin marks at the silent AG locus. Simultaneous loss of ATX1 and CLF restored AG repression and normalized leaf phenotypes. At the molecular level, disrupted ATX1 and CLF functions did not lead to erasure of the CLF- and ATX1-generated epigenetic marks, as expected: instead, in the double mutants, H3K27me3 and H3K4me3 tags were partially restored. We demonstrate that ATX1 and CLF physically interact linking mechanistically the observed effects.

BackgroundChanges in gene expression enable organisms to respond to environmental stress. Levels of cellular lipid second messengers, such as the phosphoinositide PtdIns5P, change in response to a variety of stresses and can modulate the localization, conformation and activity of a number of intracellular proteins. The plant trithorax factor (ATX1) tri-methylates the lysine 4 residue of histone H3 (H3K4me3) at gene coding sequences, which positively correlates with gene transcription. Microarray analysis has identified a target gene (WRKY70) that is regulated by both ATX1 and by the exogenous addition of PtdIns5P in Arabidopsis. Interestingly, ATX1 contains a PtdIns5P interaction domain (PHD finger) and thus, phosphoinositide signaling, may link environmental stress to changes in gene transcription.Principal FindingsUsing the plant Arabidopsis as a model system, we demonstrate a link between PtdIns5P and the activity of the chromatin modifier ATX1 in response to dehydration stress. We show for the first time that dehydration leads to an increase in cellular PtdIns5P in Arabidopsis. The Arabidopsis homolog of myotubularin (AtMTM1) is capable of generating PtdIns5P and here, we show that AtMTM1 is essential for the induced increase in PtdIns5P upon dehydration. Furthermore, we demonstrate that the ATX1-dependent gene, WRKY70, is downregulated during dehydration and that lowered transcript levels are accompanied by a drastic reduction in H3K4me3 of its nucleosomes. We follow changes in WRKY70 nucleosomal K4 methylation as a model to study ATX1 activity at chromatin during dehydration stress. We found that during dehydration stress, the physical presence of ATX1 at the WRKY70 locus was diminished and that ATX1 depletion resulted from it being retained in the cytoplasm when PtdIns5P was elevated. The PHD of ATX1 and catalytically active AtMTM1 are required for the cytoplasmic localization of ATX1.Conclusions/SignificanceThe novelty of the manuscript is in the discovery of a mechanistic link between a chromatin modifying activity (ATX1) and a lipid (PtdIns5P) synthesis in a signaling pathway that ultimately results in altered expression of ATX1 dependent genes downregulated in response to dehydration stress.