Oryza sativa RNA-Dependent RNA Polymerase 6 Contributes to Double-Strand Break Formation in Meiosis.

RNA-dependent RNA polymerase 6 (RDR6) is a core component of the small RNA biogenesis pathway, but its function in meiosis is unclear. Here, we report a new allele of OsRDR6 (Osrdr6-meiosis [Osrdr6-mei]), which causes meiosis-specific phenotypes in rice (Oryza sativa). In Osrdr6-mei, meiotic double-strand break (DSB) formation is partially blocked. We created a biallelic mutant with more severe phenotypes, Osrdr6-bi, by crossing Osrdr6-mei with a knockout mutant, Osrdr6-edit In Osrdr6-bi meiocytes, 24 univalents were observed, and no histone H2AX phosphorylation foci were detected. Compared with the wild type, the number of 21-nucleotide small RNAs in Osrdr6-mei was dramatically lower, while the number of 24-nucleotide small RNAs was significantly higher. Thousands of differentially methylated regions (DMRs) were discovered in Osrdr6-mei, implying that OsRDR6 plays an important role in DNA methylation. There were 457 genes downregulated in Osrdr6-mei, including three genes, CENTRAL REGION COMPONENT1, P31 comet , and O. sativa SOLO DANCERS, related to DSB formation. Interestingly, the downregulated genes were associated with a high level of 24-nucleotide small RNAs but less strongly associated with DMRs. Therefore, we speculate that the alteration in expression of small RNAs in Osrdr6 mutants leads to the defects in DSB formation during meiosis, which might not be directly dependent on RNA-directed DNA methylation.

miR1432-OsACOT (Acyl-CoA thioesterase) module determines grain yield via enhancing grain filling rate in rice.

Rice grain filling rate contributes largely to grain productivity and accumulation of nutrients. MicroRNAs (miRNAs) are key regulators of development and physiology in plants and become a novel key target for engineering grain size and crop yield. However, there is little studies, so far, showing the miRNA regulation of grain filling and rice yield, in consequence. Here, we show that suppressed expression of rice miR1432 (STTM1432) significantly improves grain weight by enhancing grain filling rate and leads to an increase in overall grain yield up to 17.14% in a field trial. Molecular analysis identified rice Acyl-CoA thioesterase (OsACOT), which is conserved with ACOT13 in other species, as a major target of miR1432 by cleavage. Moreover, overexpression of miR1432-resistant form of OsACOT (OXmACOT) resembled the STTM1432 plants, that is, a large margin of an increase in grain weight up to 46.69% through improving the grain filling rate. Further study indicated that OsACOT was involved in biosynthesis of medium-chain fatty acids. In addition, RNA-seq based transcriptomic analyses of transgenic plants with altered expression of miR1432 demonstrated that downstream genes of miR1432-regulated network are involved in fatty acid metabolism and phytohormones biosynthesis and also overlap with the enrichment analysis of co-expressed genes of OsACOT, which is consistent with the increased levels of auxin and abscisic acid in STTM1432 and OXmACOT plants. Overall, miR1432-OsACOT module plays an important role in grain filling in rice, illustrating its capacity for engineering yield improvement in crops.

Suppression of microRNA159 impacts multiple agronomic traits in rice (Oryza sativa L.).

BACKGROUND: microRNAs (miRNAs) are important regulators in plant growth and development. miR159 is a conserved miRNA among different plant species and has various functions in plants. Studies on miR159 are mostly done on model plant, Arabidopsis thaliana. In rice, studies on miR159 were either based upon genome-wide expression analyses focused upon responses to different nitrogen forms and abiotic stress or upon phenotypic studies of transgenic plants overexpressing its precursor. STTM (Short Tandem Target Mimic) is an effective tool to block the activity of endogenous mature miRNA activity in plant. Therefore, specific roles of miR159 in rice could be explored by down regulating miR159 through STTM. RESULTS: In this study, expression of mature miR159 was successfully suppressed by STTM which resulted in the increased expressions of its two targets genes, OsGAMYB and OsGAMYBL1 (GAMYB-LIKE 1). Overall, STTM159 plants exhibited short stature along with smaller organ size and reduction in stem diameter, length of flag leaf, main panicle, spikelet hulls and grain size. Histological analysis of stem, leaf and mature spikelet hull showed the reduced number of small vascular bundles (SVB), less number of small veins (SV) between two big veins (LV) and less cell number in outer parenchyma. Gene Ontology (GO) enrichment analysis of differentially expressed genes between wild type plants and STTM159 transgenic plants showed that genes involved in cell division, auxin, cytokinin (CK) and brassinosteroids (BRs) biosynthesis and signaling are significantly down-regulated in STTM159 plants. CONCLUSION: Our data suggests that in rice, miR159 positively regulates organ size, including stem, leaf, and grain size due to the promotion of cell division. Further analysis from the RNA-seq data showed that the decreased cell divisions in STTM159 transgenic plants may result, at least partly from the lower expression of the genes involved in cell cycle and hormone homeostasis, which provides new insights of rice miR159-specific functions.

Bacillus cereus NJ01 induces SA- and ABA-mediated immunity against bacterial pathogens through the EDS1-WRKY18 module.

Emerging evidence suggests a beneficial role of rhizobacteria in ameliorating plant disease resistance in an environment-friendly way. In this study, we characterize a rhizobacterium, Bacillus cereus NJ01, that enhances bacterial pathogen resistance in rice and Arabidopsis. Transcriptome analyses show that root inoculation of NJ01 induces the expression of salicylic acid (SA)- and abscisic acid (ABA)-related genes in Arabidopsis leaves. Genetic evidence showed that EDS1, PAD4, and WRKY18 are required for B. cereus NJ01-induced bacterial resistance. An EDS1-PAD4 complex interacts with WRKY18 and enhances its DNA binding activity. WRKY18 directly binds to the W box in the promoter region of the SA biosynthesis gene ICS1 and ABA biosynthesis genes NCED3 and NCED5 and contributes to the NJ01-induced bacterial resistance. Taken together, our findings indicate a role of the EDS1/PAD4-WRKY18 complex in rhizobacteria-induced disease resistance.

Shengjiang San alleviated sepsis-induced lung injury through its bidirectional regulatory effect.

BACKGROUND: Sepsis is a life-threatening organ dysfunction caused by dysregulated host responses to infection, for which effective therapeutic strategies are still absent. Shengjiang San (SJS), a well-known Traditional Chinese Medicine formula, has been widely used clinically. However, its role in sepsis-induced lung injury remains unclear. METHODS: To explore its specific mechanism, we firstly established a sepsis animal model using cecal ligation and puncture (CLP) and treated MH-S cells with LPS plus ATP. Then, UPLC/Q-TOF-MS/MS was utilized to identify its active ingredients. Network pharmacology analysis was performed to uncover the potential mechanism. HE staining and biochemical analysis were conducted to validate its therapeutic effect. ELISA was applied to detect the release of pro-inflammatory and anti-inflammatory cytokines. Western blot was utilized to detect the protein levels of GSDMD, NLRP3, P65, ASC and caspase-1. RESULTS: SJS could dramatically increase the survival rate of sepsis. In addition, it is able to inhibit the pro-inflammatory cytokines release at day 1 post CLP while promote their production at day 7, indicating SJS could attenuate uncontrolled inflammatory response in the early stage and improve immunosuppression in the late phase. Network pharmacology analysis showed that pyroptosis is the crucial action SJS exerted in the protection of sepsis-induced lung injury. Western blot data implicated SJS could attenuate pyroptosis in early sepsis while enhance in the late phase. CONCLUSIONS: SJS acted to alleviate sepsis-induced lung injury through its bidirectional regulatory effect.

High temperature and drought stress cause abscisic acid and reactive oxygen species accumulation and suppress seed germination growth in rice.

Seed germination is one of the most important biological processes in the life cycle of plants, and temperature and water are the two most critical environmental factors that influence seed germination. In the present study, we investigated the roles of the plant hormone abscisic acid (ABA) and reactive oxygen species (ROS) in high temperature (HT) and drought-induced inhibition of rice seed germination. HT and drought stress caused ABA accumulation in seeds and inhibited seed germination and seedling establishment. Quantitative real-time polymerase chain reaction analysis revealed that HT and drought stress induced the expression of OsNCED3, a key gene in ABA synthesis in rice seeds. In addition, ROS (O2•- and H2O2) and malondialdehyde contents were increased in germinating seeds under HT and drought stress. Moreover, we adopted the non-invasive micro-test technique to detect H2O2 and Ca2+ fluxes at the site of coleoptile emergence. HT and drought stress resulted in a H2O2 efflux, but only drought stress significantly induced Ca2+ influx. Antioxidant enzyme assays revealed that superoxide dismutase (SOD), peroxidase, catalase (CAT), and ascorbate peroxidase (APX) activity were reduced by HT and drought stress, consistent with the expression of OsCu/ZnSOD, OsCATc, and OsAPX2 during seed germination. Altogether, these results suggest that ABA and ROS accumulation under HT and drought conditions can inhibit rice seed germination and growth.