Regulation of reactive oxygen species during plant immunity through phosphorylation and ubiquitination of RBOHD.

Production of reactive oxygen species (ROS) is critical for successful activation of immune responses against pathogen infection. The plant NADPH oxidase RBOHD is a primary player in ROS production during innate immunity. However, how RBOHD is negatively regulated remains elusive. Here we show that RBOHD is regulated by C-terminal phosphorylation and ubiquitination. Genetic and biochemical analyses reveal that the PBL13 receptor-like cytoplasmic kinase phosphorylates RBOHD's C-terminus and two phosphorylated residues (S862 and T912) affect RBOHD activity and stability, respectively. Using protein array technology, we identified an E3 ubiquitin ligase PIRE (PBL13 interacting RING domain E3 ligase) that interacts with both PBL13 and RBOHD. Mimicking phosphorylation of RBOHD (T912D) results in enhanced ubiquitination and decreased protein abundance. PIRE and PBL13 mutants display higher RBOHD protein accumulation, increased ROS production, and are more resistant to bacterial infection. Thus, our study reveals an intricate post-translational network that negatively regulates the abundance of a conserved NADPH oxidase

Stromule extension along microtubules coordinated with actin-mediated anchoring guides perinuclear chloroplast movement during innate immunity.

Dynamic tubular extensions from chloroplasts called stromules have recently been shown to connect with nuclei and function during innate immunity. We demonstrate that stromules extend along microtubules (MTs) and MT organization directly affects stromule dynamics since stabilization of MTs chemically or genetically increases stromule numbers and length. Although actin filaments (AFs) are not required for stromule extension, they provide anchor points for stromules. Interestingly, there is a strong correlation between the direction of stromules from chloroplasts and the direction of chloroplast movement. Stromule-directed chloroplast movement was observed in steady-state conditions without immune induction, suggesting it is a general function of stromules in epidermal cells. Our results show that MTs and AFs may facilitate perinuclear clustering of chloroplasts during an innate immune response. We propose a model in which stromules extend along MTs and connect to AF anchor points surrounding nuclei, facilitating stromule-directed movement of chloroplasts to nuclei during innate immunity

The plastidial retrograde signal methyl erythritol cyclopyrophosphate is a regulator of salicylic acid and jasmonic acid crosstalk.

The exquisite harmony between hormones and their corresponding signaling pathways is central to prioritizing plant responses to simultaneous and/or successive environmental trepidations. The crosstalk between jasmonic acid (JA) and salicylic acid (SA) is an established effective mechanism that optimizes and tailors plant adaptive responses. However, the underlying regulatory modules of this crosstalk are largely unknown. Global transcriptomic analyses of mutant plants (ceh1) with elevated levels of the stress-induced plastidial retrograde signaling metabolite 2-C-methyl-D-erythritol cyclopyrophosphate (MEcPP) revealed robustly induced JA marker genes, expected to be suppressed by the presence of constitutively high SA levels in the mutant background. Analyses of a range of genotypes with varying SA and MEcPP levels established the selective role of MEcPP-mediated signal(s) in induction of JA-responsive genes in the presence of elevated SA. Metabolic profiling revealed the presence of high levels of the JA precursor 12-oxo-phytodienoic acid (OPDA), but near wild type levels of JA in the ceh1 mutant plants. Analyses of coronatine-insensitive 1 (coi1)/ceh1 double mutant plants confirmed that the MEcPP-mediated induction is JA receptor COI1 dependent, potentially through elevated OPDA. These findings identify MEcPP as a previously unrecognized central regulatory module that induces JA-responsive genes in the presence of high SA, thereby staging a multifaceted plant response within the environmental context

Virus-induced gene silencing database for phenomics and functional genomics in Nicotiana benthamiana

Virus-induced gene silencing (VIGS) is an important forward and reverse genetics method for the study of gene function in many plant species, especially Nicotiana benthamiana. However, despite the widespread use of VIGS, a searchable database compiling the phenotypes observed with this method is lacking. Such a database would allow researchers to know the phenotype associated with the silencing of a large number of individual genes without experimentation. We have developed a VIGS phenomics and functional genomics database (VPGD) that has DNA sequence information derived from over 4,000 N. benthamiana VIGS clones along with the associated silencing phenotype for approximately 1,300 genes. The VPGD has a built-in BLAST search feature that provides silencing phenotype information of specific genes. In addition, a keyword-based search function could be used to find a specific phenotype of interest with the corresponding gene, including its Gene Ontology descriptions. Query gene sequences from other plant species that have not been used for VIGS can also be searched for their homologs and silencing phenotype in N. benthamiana. VPGD is useful for identifying gene function not only in N. benthamiana but also in related Solanaceae plants such as tomato and potato. The database is accessible at http://vigs.noble.org.Noble Research Institute and NSF IOS-102564

SCREENING OF PHYTOCHEMICAL CONSTITUENTS OF THE LEAVES OF CLINACANTHUS SIAMENSIS BREMEK AND CISSAMPELOS PAREIRA L USED AS ANTIDOTE FOR SNAKE BITE IN INDIGENOUS MEDICINE

Objective: The study is aimed to screen the phytochemical constituents of the medicinal plants Clinacanthus siamensis Bremek and Cissampelos pareira L used as anti snake venom in tribal medicine. Methods: The phytochemical compounds such as proteins, carbohydrates, lipids, phenols, tannins, flavonoids, saponins, steroids, terpenoids, coumarins, anthocyanin, glycosides, reducing sugars, lignins, anthoquinone and alkaloids were extracted by standard methods and compared the components for its antivenom activity. Results: Qualitative analysis of methanolic and aqueous extracts of leaves confirmed the presence of primary metabolites like proteins, carbohydrates, lipids and reducing sugars and secondary metabolites like phenols, tannins, flavonoids, glycosides, saponins, steroids, terpenoids, coumarins, lignins, anthocyanin, anthoquinone and alkaloids. Quantitative estimation of primary and secondary metabolites showed that the presence of proteins, flavonoids, tannins, alkaloids and lipids are more in Cissampelos pareira L when compared to Clinacanthus siamensis B. whereas the phenols and carbohydrates were more in Clinacanthus siamensis B. Conclusion: The study helped in the successful screening of phytochemical constituents which supports the traditional knowledge of the use of the plants as important medicine, as an antidote for poisonous snake bites and in curing various ailments

Antioxidant activity and Cytotoxic evaluation of Phytofabricated Silver Nanoparticles of Fig (Ficus mollis Vahl)

The present study aimed to evaluate Antioxidant and Cytotoxic activity of phytofabricated silver nanoparticles (FmF-AgNPs) derived from Figs of Ficus mollis. This green synthesized FmF-AgNPs were tested for antioxidant activity with DPPH assay and cytotoxicity activity against MCF-7 (Human breast adenocarcinoma cell lines) with MTT assay at various concentrations. The data obtained demonstrated that FmF-AgNPs posses both antioxidant activity and cytotoxicity activity which is dosage-dependent. In conclusion, results obtained revealed the potent therapeutic value of phytofabricated silver nanoparticles (FmF-AgNPs) can act as potent antioxidant and anticancer agent