WRKY Transcription Factors Involved in Activation of SA Biosynthesis Genes

<p>Abstract</p> <p>Background</p> <p>Increased defense against a variety of pathogens in plants is achieved through activation of a mechanism known as systemic acquired resistance (SAR). The broad-spectrum resistance brought about by SAR is mediated through salicylic acid (SA). An important step in SA biosynthesis in Arabidopsis is the conversion of chorismate to isochorismate through the action of isochorismate synthase, encoded by the <it>ICS1 </it>gene. Also <it>AVR</it>_{<it>PPHB </it>}<it>SUSCEPTIBLE 3 </it>(<it>PBS3</it>) plays an important role in SA metabolism, as <it>pbs3 </it>mutants accumulate drastically reduced levels of SA-glucoside, a putative storage form of SA. Bioinformatics analysis previously performed by us identified WRKY28 and WRKY46 as possible regulators of <it>ICS1 </it>and <it>PBS3</it>.</p> <p>Results</p> <p>Expression studies with <it>ICS1 promoter::β-glucuronidase </it>(<it>GUS</it>) genes in <it>Arabidopsis thaliana </it>protoplasts cotransfected with <it>35S::WRKY28 </it>showed that over expression of WRKY28 resulted in a strong increase in GUS expression. Moreover, qRT-PCR analyses indicated that the endogenous <it>ICS1 </it>and <it>PBS3 </it>genes were highly expressed in protoplasts overexpressing WRKY28 or WRKY46, respectively. Electrophoretic mobility shift assays indentified potential WRKY28 binding sites in the <it>ICS1 </it>promoter, positioned -445 and -460 base pairs upstream of the transcription start site. Mutation of these sites in protoplast transactivation assays showed that these binding sites are functionally important for activation of the <it>ICS1 </it>promoter. Chromatin immunoprecipitation assays with haemagglutinin-epitope-tagged WRKY28 showed that the region of the <it>ICS1 </it>promoter containing the binding sites at -445 and -460 was highly enriched in the immunoprecipitated DNA.</p> <p>Conclusions</p> <p>The results obtained here confirm results from our multiple microarray co-expression analyses indicating that WRKY28 and WRKY46 are transcriptional activators of <it>ICS1 </it>and <it>PBS3</it>, respectively, and support this <it>in silico </it>screening as a powerful tool for identifying new components of stress signaling pathways.</p

Prospecting for Genes involved in transcriptional regulation of plant defenses, a bioinformatics approach

<p>Abstract</p> <p>Background</p> <p>In order to comprehend the mechanisms of induced plant defense, knowledge of the biosynthesis and signaling pathways mediated by salicylic acid (SA), jasmonic acid (JA) and ethylene (ET) is essential. Potentially, many transcription factors could be involved in the regulation of these pathways, although finding them is a difficult endeavor. Here we report the use of publicly available Arabidopsis microarray datasets to generate gene co-expression networks.</p> <p>Results</p> <p>Using 372 publicly available microarray data sets, a network was constructed in which Arabidopsis genes for known components of SA, JA and ET pathways together with the genes of over 1400 transcription factors were assayed for co-expression. After determining the Pearson Correlation Coefficient cutoff to obtain the most probable biologically relevant co-expressed genes, the resulting network confirmed the presence of many genes previously reported in literature to be relevant for stress responses and connections that fit current models of stress gene regulation, indicating the potential of our approach. In addition, the derived network suggested new candidate genes and associations that are potentially interesting for future research to further unravel their involvement in responses to stress.</p> <p>Conclusions</p> <p>In this study large sets of stress related microarrays were used to reveal co-expression networks of transcription factors and signaling pathway components. These networks will benefit further characterization of the signal transduction pathways involved in plant defense.</p

Evolutionary divergence of the plant elicitor peptides (Peps) and their receptors: interfamily incompatibility of perception but compatibility of downstream signalling

Plant elicitor peptides (Peps) co-evolved with their receptors, resulting in interfamily incompatibility of Pep recognition. In contrast, operation of defence pathways by Pep receptors is conserved within the flowering plant

HISTONE DEACETYLASE 9 stimulates auxin-dependent thermomorphogenesis in Arabidopsis thaliana by mediating H2A.Z depletion

Many plant species respond to unfavorable high ambient temperatures by adjusting their vegetative body plan to facilitate cooling. This process is known as thermomorphogenesis and is induced by the phytohormone auxin. Here, we demonstrate that the chromatin-modifying enzyme HISTONE DEACETYLASE 9 (HDA9) mediates thermomorphogenesis but does not interfere with hypocotyl elongation during shade avoidance. HDA9 is stabilized in response to high temperature and mediates histone deacetylation at the YUCCA8 locus, a rate-limiting enzyme in auxin biosynthesis, at warm temperatures. We show that HDA9 permits net eviction of the H2A.Z histone variant from nucleosomes associated with YUCCA8, allowing binding and transcriptional activation by PHYTOCHROME INTERACTING FACTOR 4, followed by auxin accumulation and thermomorphogenesis

Architecture and dynamics of the jasmonic acid gene regulatory network

Jasmonic acid (JA) is a critical hormonal regulator of plant growth and defense. To advance our understanding of the architecture and dynamic regulation of the JA gene regulatory network, we performed a high-resolution RNA-seq time series of methyl JA-treated Arabidopsis thaliana at 15 time points over a 16-h period. Computational analysis showed that methyl JA (MeJA) induces a burst of transcriptional activity, generating diverse expression patterns over time that partition into distinct sectors of the JA response targeting specific biological processes. The presence of transcription factor (TF) DNA binding motifs correlated with specific TF activity during temporal MeJA-induced transcriptional reprogramming. Insight into the underlying dynamic transcriptional regulation mechanisms was captured in a chronological model of the JA gene regulatory network. Several TFs, including MYB59 and bHLH27, were uncovered as early network components with a role in pathogen and insect resistance. Analysis of subnetworks surrounding the TFs ORA47, RAP2.6L, MYB59, and ANAC055, using transcriptome profiling of overexpressors and mutants, provided insights into their regulatory role in defined modules of the JA network. Collectively, our work illuminates the complexity of the JA gene regulatory network, pinpoints and validates previously unknown regulators, and provides a valuable resource for functional studies on JA signaling components in plant defense and development

Towards predicting the environmental metabolome from metagenomics with a mechanistic model

The environmental metabolome and metabolic potential of microorganisms are dominant and essential factors shaping microbial community composition. Recent advances in genome annotation and systems biology now allow us to semiautomatically reconstruct genome-scale metabolic models (GSMMs) of microorganisms based on their genome sequence 1 . Next, growth of these models in a defined metabolic environment can be predicted in silico, mechanistically linking the metabolic fluxes of individual microbial populations to the community dynamics. A major advantage of GSMMs is that no training data is needed, besides information about the metabolic capacity of individual genes (genome annotation) and knowledge of the available environmental metabolites that allow the microorganism to grow. However, the composition of the environment is often not fully determined and remains difficult to measure 2 . We hypothesized that the relative abundance of different bacterial species, as measured by metagenomics, can be combined with GSMMs of individual bacteria to reveal the metabolic status of a given biome. Using a newly developed algorithm involving over 1,500 GSMMs of human-associated bacteria, we inferred distinct metabolomes for four human body sites that are consistent with experimental data. Together, we link the metagenome to the metabolome in a mechanistic framework towards predictive microbiome modelling

A Novel WRKY Transcription Factor Is Required for Induction of PR-1a Gene Expression by Salicylic Acid and Bacterial Elicitors[C][W]

PR-1a is a salicylic acid-inducible defense gene of tobacco (Nicotiana tabacum). One-hybrid screens identified a novel tobacco WRKY transcription factor (NtWRKY12) with specific binding sites in the PR-1a promoter at positions −564 (box WK1) and −859 (box WK2). NtWRKY12 belongs to the class of transcription factors in which the WRKY sequence is followed by a GKK rather than a GQK sequence. The binding sequence of NtWRKY12 (WK box TTTTCCAC) deviated significantly from the consensus sequence (W box TTGAC[C/T]) shown to be recognized by WRKY factors with the GQK sequence. Mutation of the GKK sequence in NtWRKY12 into GQK or GEK abolished binding to the WK box. The WK1 box is in close proximity to binding sites in the PR-1a promoter for transcription factors TGA1a (as-1 box) and Myb1 (MBSII box). Expression studies with PR-1a promoter∷β-glucuronidase (GUS) genes in stably and transiently transformed tobacco indicated that NtWRKY12 and TGA1a act synergistically in PR-1a expression induced by salicylic acid and bacterial elicitors. Cotransfection of Arabidopsis thaliana protoplasts with 35S∷NtWRKY12 and PR-1a∷GUS promoter fusions showed that overexpression of NtWRKY12 resulted in a strong increase in GUS expression, which required functional WK boxes in the PR-1a promoter

Root transcriptional dynamics induced by beneficial rhizobacteria and microbial immune elicitors reveal signatures of adaptation to mutualists

Below ground, microbe-associated molecular patterns (MAMPs) of root-associated microbiota can trigger costly defenses at the expense of plant growth. However, beneficial rhizobacteria, such as Pseudomonas simiae WCS417, promote plant growth and induce systemic resistance without being warded off by local root immune responses. To investigate early root responses that facilitate WCS417 to exert its plant-beneficial functions, we performed time series RNA-Seq of Arabidopsis roots in response to live WCS417 and compared it with MAMPs flg22(417) (from WCS417), flg22(Pa) (from pathogenic Pseudomonas aeruginosa) and fungal chitin. The MAMP transcriptional responses differed in timing, but displayed a large overlap in gene identity. MAMP-upregulated genes are enriched for genes with functions in immunity, while downregulated genes are enriched for genes related to growth and development. Although 74% of the transcriptional changes inflicted by live WCS417 overlapped with the flg22(417) profile, WCS417 actively suppressed more than half of the MAMP-triggered transcriptional responses, possibly to allow the establishment of a mutually beneficial interaction with the host root. Interestingly, the sector of the flg22(417) -repressed transcriptional network that is not affected by WCS417 has a strong auxin signature. Using auxin response mutant tir1afb2afb3, we demonstrate a dual role for auxin signaling in finely balancing growth-promoting and defense-eliciting activities of beneficial microbes in plant roots