Project description:Brassinosteroid (BR) and auxin co-regulate plant growth in a process termed cross-talking. Based on the assumption that their signal transductions are partially shared, inhibitory chemicals for both signal transductions were screened from a commercially-available library. A chemical designated as NJ15 (ethyl 2-[5-(3,5-dichlorophenyl)-1,2,3,4-tetrazole-2-yl]acetate) diminished the growth promotion of both adzuki bean epicotyls and Arabidopsis seedlings, by either the application of BR or auxin. To understand its target site(s), bioassays with a high dependence on either the signal transduction of BR (BR-signaling) or of auxin (AX-signaling), were performed. NJ15 inhibited photomorphogenesis of Arabidopsis seedlings grown in the dark, which mainly depends on BR-signaling, while NJ15 also inhibited their gravitropic responses mainly depending on AX-signaling. On the study for the structure-activity relationships of NJ15 analogues, they showed strong correlations on the inhibitory profiles between BR- and AX-signalings. These correlations imply that NJ15 targets the downstream pathway after the integration of BR- and AX-signals.
Project description:Brassinosteroids (BRs) are a class of class of phytohormones with important roles in regulating physiological and developmental processes. Small RNAs, including small interfering RNAs and microRNAs (miRNAs), are non-protein coding RNAs that regulate gene expression at the transcriptional and post-transcriptional levels. However, the roles of small RNAs in BR response have not been studied well. In this study, we aimed to identify BR-responsive small RNA clusters and miRNAs in Arabidopsis. In addition, the effect of BR-responsive small RNAs on their transcripts and target genes were examined. Small RNA libraries were constructed from control and epibrassinolide-treated seedlings. After sequencing the small RNA libraries, differentially expressed small RNA clusters were identified by examining the expression levels of small RNAs in 100-nt bins of Arabidopsis genome. To identify the BR-responsive miRNAs, the expression levels of all the annotated mature miRNAs, registered in miRBase, were analyzed. Previously published RNA-seq data were utilized to monitor the BR-responsive expression patterns of differentially expressed small RNA clusters and miRNA target genes. In results, 38 BR-responsive small RNA clusters, including 30 down-regulated and eight up-regulated clusters, were identified. These differentially expressed small RNA clusters were from miRNA loci, transposons, protein-coding genes, pseudo genes and others. Of these, a transgene, BRI1, accumulates small RNAs, which are not found in the wild type. Small RNAs in this transgene are up-regulated by BRs while BRI1 mRNA is down-regulated by BRs. By analyzing the expression patterns of mature miRNAs, we have identified BR-repressed miR398a-5p and BR-induced miR156g. Although miR398a-5p is down-regulated by BRs, its predicted targets were not responsive to BRs. However, SPL3, a target of BR-inducible miR156g, is down-regulated by BRs. BR-responsive small RNAs and miRNAs identified in this study will provide an insight into the role of small RNAs in BR responses in plants. Especially, we suggest that miR156g/SPL3 module might play a role in BR-mediated growth and development in Arabidopsis.
Project description:Brassinosteroid (BR) and auxin co-regulate plant growth in a process termed cross-talking. Based on the assumption that their signal transductions are partially shared, inhibitory chemicals for both signal transductions were screened from a commercially-available library. A chemical designated as NJ15 (ethyl 2-[5-(3,5-dichlorophenyl)-1,2,3,4-tetrazole-2-yl]acetate) diminished the growth promotion of both adzuki bean epicotyls and Arabidopsis seedlings, by either the application of BR or auxin. To understand its target site(s), bioassays with a high dependence on either the signal transduction of BR (BR-signaling) or of auxin (AX-signaling), were performed. NJ15 inhibited photomorphogenesis of Arabidopsis seedlings grown in the dark, which mainly depends on BR-signaling, while NJ15 also inhibited their gravitropic responses mainly depending on AX-signaling. On the study for the structure-activity relationships of NJ15 analogues, they showed strong correlations on the inhibitory profiles between BR- and AX-signalings. These correlations imply that NJ15 targets the downstream pathway after the integration of BR- and AX-signals. Arabidopsis thaliana ecotype Col-0 was used as the wild type (WT) in this study. The Col-0 plants, grown vertically as a gravitropic response assay for 4 days on MS plates, were sprayed with mock or with 10 µM NJ15 (ethyl 2-[5-(3,5-dichlorophenyl)-1,2,3,4-tetrazole-2-yl]acetate) and were gravistimulated by turning then 90° relative to the first direction. After 8 h, plants were harvested and total RNA was extracted with a Total RNA Extraction Kit Mini for plant (RBC Bioscience), then total RNA was subjected to microarray analysis.