SUB1A-1 anchors a regulatory cascade for epigenetic and transcriptional controls of submergence tolerance in rice.

Most rice (Oryza sativa) cultivars cannot survive under prolonged submergence. However, some O. sativa ssp. indica cultivars, such as FR13A, are highly tolerant owing to the SUBMERGENCE 1A-1 (SUB1A-1) allele, which encodes a Group VII ethylene-responsive factor (ERFVII) protein; other submergence-intolerant cultivars contain a SUB1A-2 allele. The two alleles differ only by a single substitution at the 186th amino acid position from serine in SUB1A-1 to proline in SUB1A-2 resulting in only SUB1A-1 being able to be phosphorylated. Two other ERFVIIs, ERF66 and ERF67, function downstream of SUB1A-1 to form a regulatory cascade in response to submergence stress. Here, we show that SUB1A-1, but not SUB1A-2, interacts with ADA2b of the ADA2b-GCN5 acetyltransferase complex, in which GCN5 functions as a histone acetyltransferase. Phosphorylation of SUB1A-1 at serine 186 enhances the interaction of SUB1A-1 with ADA2b. ADA2b and GCN5 expression was induced under submergence, suggesting that these two genes might play roles in response to submergence stress. In transient assays, binding of SUB1A-1 to the ERF67 promoter and ERF67 transcription were highly induced when SUB1A-1 was expressed together with the ADA2b-GCN5 acetyltransferase complex. Taken together, these results suggest that phospho-SUB1A-1 recruits the ADA2-GCN5 acetyltransferase complex to modify the chromatin structure of the ERF66/ERF67 promoter regions and activate gene expression, which in turn enhances rice submergence tolerance.

Ethylene modulates translation dynamics in Arabidopsis under submergence via GCN2 and EIN2.

General translational repression is a key process that reduces energy consumption under hypoxia. Here, we show that plant stress-activated general control nonderepressible 2 (GCN2) was activated to regulate the reduction in polysome loading during submergence in Arabidopsis. GCN2 signaling was activated by ethylene under submergence. GCN2 activity was reduced in etr1-1, but not in ein2-5 or eil1ein3, under submergence, suggesting that GCN2 activity is regulated by a noncanonical ethylene signaling pathway. Polysome loading was not reduced in ein2-5 under submergence, implying that ethylene modulates translation via both EIN2 and GCN2. Transcriptomic analysis demonstrated that EIN2 and GCN2 regulate not only general translational repression but also translational enhancement of specific mRNAs under submergence. Together, these results demonstrate that during submergence, entrapped ethylene triggers GCN2 and EIN2 to regulate translation dynamics and ensure the translation of stress response proteins.

Regulatory cascade involving transcriptional and N-end rule pathways in rice under submergence.

The rice SUB1A-1 gene, which encodes a group VII ethylene response factor (ERFVII), plays a pivotal role in rice survival under flooding stress, as well as other abiotic stresses. In Arabidopsis, five ERFVII factors play roles in regulating hypoxic responses. A characteristic feature of Arabidopsis ERFVIIs is a destabilizing N terminus, which functions as an N-degron that targets them for degradation via the oxygen-dependent N-end rule pathway of proteolysis, but permits their stabilization during hypoxia for hypoxia-responsive signaling. Despite having the canonical N-degron sequence, SUB1A-1 is not under N-end rule regulation, suggesting a distinct hypoxia signaling pathway in rice during submergence. Herein we show that two other rice ERFVIIs gene, ERF66 and ERF67, are directly transcriptionally up-regulated by SUB1A-1 under submergence. In contrast to SUB1A-1, ERF66 and ERF67 are substrates of the N-end rule pathway that are stabilized under hypoxia and may be responsible for triggering a stronger transcriptional response to promote submergence survival. In support of this, overexpression of ERF66 or ERF67 leads to activation of anaerobic survival genes and enhanced submergence tolerance. Furthermore, by using structural and protein-interaction analyses, we show that the C terminus of SUB1A-1 prevents its degradation via the N-end rule and directly interacts with the SUB1A-1 N terminus, which may explain the enhanced stability of SUB1A-1 despite bearing an N-degron sequence. In summary, our results suggest that SUB1A-1, ERF66, and ERF67 form a regulatory cascade involving transcriptional and N-end rule control, which allows rice to distinguish flooding from other SUB1A-1-regulated stresses.

Analysis of genetic diversity of Xanthomonas oryzae pv. oryzae populations in Taiwan.

Rice bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo) is a major rice disease. In Taiwan, the tropical indica type of Oryza sativa originally grown in this area is mix-cultivated with the temperate japonica type of O. sativa, and this might have led to adaptive changes of both rice host and Xoo isolates. In order to better understand how Xoo adapts to this unique environment, we collected and analyzed fifty-one Xoo isolates in Taiwan. Three different genetic marker systems consistently identified five groups. Among these groups, two of them had unique sequences in the last acquired ten spacers in the clustered regularly interspaced short palindromic repeats (CRISPR) region, and the other two had sequences that were similar to the Japanese isolate MAFF311018 and the Philippines isolate PXO563, respectively. The genomes of two Taiwanese isolates with unique CRISPR sequence features, XF89b and XM9, were further completely sequenced. Comparison of the genome sequences suggested that XF89b is phylogenetically close to MAFF311018, and XM9 is close to PXO563. Here, documentation of the diversity of groups of Xoo in Taiwan provides evidence of the populations from different sources and hitherto missing information regarding distribution of Xoo populations in East Asia.

Submergence confers immunity mediated by the WRKY22 transcription factor in Arabidopsis.

Transcriptional control plays an important role in regulating submergence responses in plants. Although numerous genes are highly induced during hypoxia, their individual roles in hypoxic responses are still poorly understood. Here, we found that expression of genes that encode members of the WRKY transcription factor family was rapidly and strongly induced upon submergence in Arabidopsis thaliana, and this induction correlated with induction of a large portion of innate immunity marker genes. Furthermore, prior submergence treatment conferred higher resistance to the bacterial pathogen Pseudomonas syringae in Arabidopsis. Among the WRKY genes tested, WRKY22 had the highest level of induction during the early stages of submergence. Compared with the wild type, WRKY22 T-DNA insertion mutants wrky22-1 and wrky22-2 had lower disease resistance and lower induction of innate immunity markers, such as FLG22-INDUCED RECEPTOR-LIKE KINASE1 (FRK1) and WRKY53, after submergence. Furthermore, transcriptomic analyses of wrky22-2 and chromatin immunoprecipitation identified several potential targets of WRKY22, which included genes encoding a TIR domain-containing protein, a plant peptide hormone, and many OLIGO PEPTIDE TRANSPORTER genes, all of which may lead to induction of innate immunity. In conclusion, we propose that submergence triggers innate immunity in Arabidopsis via WRKY22, a response that may protect against a higher probability of pathogen infection either during or after flooding.

Insights into hypoxic systemic responses based on analyses of transcriptional regulation in Arabidopsis.

We have adopted a hypoxic treatment system in which only roots were under hypoxic conditions. Through analyzing global transcriptional changes in both shoots and roots, we found that systemic signals may be transduced from roots to trigger responses in tissues not directly subjected to hypoxia. The molecular mechanisms of such systemic responses under flooding are currently largely unknown. Using ontological categorization for regulated genes, a systemic managing program of carbohydrate metabolism was observed, providing an example of how systemic responses might facilitate the survival of plants under flooding. Moreover, a proportion of gene expressions that regulated in shoots by flooding was affected in an ethylene signaling mutation, ein2-5. Many systemic-responsive genes involved in the systemic carbohydrate managing program, hormone responses and metabolism, ubiquitin-dependent protein degradation were also affected in ein2-5. These results suggested an important role of ethylene in mediation of hypoxic systemic responses. Genes associated with abscisic acid (ABA) biosynthesis are upregulated in shoots and down regulated in roots. An ABA signaling mutation, abi4-1, affects expression of several systemic responsive genes. These results suggested that regulation of ABA biosynthesis could be required for systemic responses. The implications of these results for the systemic responses of root-flooded Arabidopsis are discussed.