Genome-wide identification and expression profiling of chitinase genes in tea (Camellia sinensis (L.) O. Kuntze) under biotic stress conditions.

Chitinases are a diverse group of enzymes having the ability to degrade chitin. Chitin is the second most abundant polysaccharide on earth, predominantly found in insect exoskeletons and fungal cell walls. In this study, we performed a genome-wide search for chitinase genes and identified a total of 49 chitinases in tea. These genes were categorized into 5 classes, where an expansion of class V chitinases has been observed in comparison to other plant species. Extensive loss of introns in 46% of the GH18 chitinases indicates that an evolutionary pressure is acting upon these genes to lose introns for rapid gene expression. The promoter upstream regions in 65% of the predicted chitinases contain methyl-jasmonate, salicylic acid and defense responsive cis-acting elements, which may further illustrate the possible role of chitinases in tea plant's defense against various pests and pathogens. Differential expression analysis revealed that transcripts of two GH19 chitinases TEA028279 and TEA019397 got upregulated during three different fungal infections in tea. While GH19 chitinase TEA031377 showed an increase in transcript abundance in the two insect infested tea tissues. Semi-quantitative RT-PCR analysis revealed that five GH19 chitinases viz. TEA018892, TEA031484, TEA28279, TEA033470 and TEA031277 showed significant increase in expression in the tea plants challenged with a biotrophic pathogen Exobasidium vexans. The study endeavours in highlighting biotic stress responsive defensive role of chitinase genes in tea.

CircRNAs responsive to winter dormancy and spring flushing conditions of tea leaf buds.

Circular RNAs (circRNAs) are important regulators of diverse biological processes of plants. However, the evolution and potential functions of circRNAs during winter dormancy and spring bud flushing of tea plant is largely unknown. Using RNA-seq data, a total of 1184 circRNAs were identified in the winter dormant and spring bud flushing leaf samples of tea plants in two different cultivars exhibiting different duration of winter dormancy. A total of 156 circRNAs are found to be differentially expressed and the weighted gene co-expression network (WGCNA) analysis revealed that 22 and 20 differentially expressed-circRNAs (DE-circRNAs) positively correlated with the flushing and dormant leaf traits, respectively, in both the tea cultivars used. Some transcription factors (TFs) viz. MYB, WRKY, ERF, bHLH and several genes related to secondary metabolite biosynthetic pathways are found to co-express with circRNAs. DE-circRNAs also predicted to interact with miRNAs and can regulate phytohormone biosynthesis and various signalling pathways in tea plant. This study uncovers the potential roles of circRNAs to determine winter dormancy and spring bud flushing conditions in tea plants.

Genome-wide identification, evolutionary relationship and expression analysis of AGO, DCL and RDR family genes in tea.

Three gene families in plants viz. Argonaute (AGOs), Dicer-like (DCLs) and RNA dependent RNA polymerase (RDRs) constitute the core components of small RNA mediated gene silencing machinery. The present study endeavours to identify members of these gene families in tea and to investigate their expression patterns in different tissues and various stress regimes. Using genome-wide analysis, we have identified 18 AGOs, 5 DCLs and 9 RDRs in tea, and analyzed their phylogenetic relationship with orthologs of Arabidopsis thaliana. Gene expression analysis revealed constitutive expression of CsAGO1 in all the studied tissues and stress conditions, whereas CsAGO10c showed most variable expression among all the genes. CsAGO10c gene was found to be upregulated in tissues undergoing high meristematic activity such as buds and roots, as well as in Exobasidium vexans infected samples. CsRDR2 and two paralogs of CsAGO4, which are known to participate in biogenesis of hc-siRNAs, showed similarities in their expression levels in most of the tea plant tissues. This report provides first ever insight into the important gene families involved in biogenesis of small RNAs in tea. The comprehensive knowledge of these small RNA biogenesis purveyors can be utilized for tea crop improvement aimed at stress tolerance and quality enhancement.

Genome wide identification and characterization of abiotic stress responsive lncRNAs in Capsicum annuum.

Long non-coding RNAs (lncRNAs) are a type of non-coding transcripts having length of more than 200 nucleotides lacking protein-coding ability. In the present study, 12807 lncRNAs were identified in Capsicum annuum tissues exposed to abiotic stress conditions viz. heat, cold, osmotic and salinity stress. Expression analysis of lncRNAs in different treatment conditions demonstrates their stress-specific expression. Thirty lncRNAs were found to act as precursors for 10 microRNAs (miRNAs) of C. annuum. Additionally, a total of 1807 lncRNAs were found to interact with 194 miRNAs which targeted 621 mRNAs of C. annuum. Among these, 344 lncRNAs were found to act as target mimics for 621 genes. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that out of those 621 gene sequences, 546 were tagged with GO terms, 105 Enzyme Code (EC) numbers were assigned to 246 genes and 223 genes are found to be involved in 63 biological pathways. In this report, we have highlighted the prospective role of lncRNAs in different abiotic stress conditions by interacting with miRNAs and regulating stress responsive transcription factors (TFs) such as DoF, WRKY, MYB, bZIP and ERF in C. annuum.

Mining of miRNAs from EST data in Dendrobium nobile.

Dendrobium nobile is an orchid species highly popular for its therapeutic properties and is often used as a medicinal herb. Documenting miRNA-target associations in D. nobile is an important step to facilitate functional genomics studies in this species. Therefore, it is of interest to identify miRNA sequences from EST data available in public databases using known techniques and tools. We report 14 potential miRNAs from three ESTs of D. nobile. They belong to 3 miRNA families (miR390, miR528 and miR414) linking to transcription factor regulation, signal transduction, DNA and protein binding, and various cellular processes covering 34 different metabolic networks in KEGG. These results help in the understanding of miRNA-mRNAs functional networks in Dendrobium nobile.

Endophytes from Gnetum gnemon L. can protect seedlings against the infection of phytopathogenic bacterium Ralstonia solanacearum as well as promote plant growth in tomato.

Endophytes are beneficial plant microbes which help the plants by producing various plant growth promoting substances and also by acting as biocontrol agents against various plant pathogens. In the present study, evaluation of endophytic bacteria isolated from Gnetum gnemon, an ethnomedicinal plant was carried out for their plant growth promoting (PGP) activity and antagonistic potential against bacterial wilt pathogen Ralstonia solanacearum. Initially a total of 40 endophytic bacteria were isolated which were clustered into 13 groups based on RFLP and BOX-PCR fingerprinting. These 13 representative isolates belonged to different genera of Bacillus (9), Solibacillus (1), Staphylococcus (2) and Caballeronia (1). Among them, six isolates were positive for production of IAA, the value of which ranged from 11.16 to 27.63 µg mL-1. Phosphate solubilisation in the range of 106.4 to -212.7 µg mL-1 was shown by three isolates. Eight isolates producedammonia, the value of which ranged from 1.3 to 6.1 µmol mL-1. All tested isolates were positive for siderophore production. For extracellular enzyme production, 7 isolates were positive for protease, 8 for cellulase and 10 for amylase production. The isolates were also tested for their antagonistic activity against R. solanacearum in vitro and in planta assay using tomato seedlings. Staphylococcus warneri GL1 showed the highest biocontrol efficacy of 77.67 % followed by Bacillus velezensis GL3 i.e. 70.1 %. R. solanacearum antagonistic isolates were analysed for the presence of antimicrobial peptide biosynthesis genes bmyB, srfAA, fenD and ituC. All the antagonistic isolates showed the presence of all four genes, except the isolate Bacillus velezensis GMC2, where the gene for fengycin synthetase (fenD) was absent. Based on in vitro PGP traits, three isolates Bacillus velezensis GL3, Bacillus atrophaeus GMC1 and Bacillus megaterium GS2 were selected, these three endophytic bacteria individually and their consortia were tested for in planta PGP activities in tomato plants. Application of Bacillus velezensis GL3 alone and consortia of three isolates showed significant improvement in growth parameters such as shoot length, fresh weight and dry weight in a pot experiment. Colonization of endosphere of treated tomato seedlings by the endophytic isolate Bacillus velezensis GL3 was confirmed by visualization of colony morphology and BOX-PCR fingerprinting. Our study highlights the potential of endophytes associated with unexplored plants like G. gnemon for development of bioformulation aimed at enhancing plant growth and bacterial wilt disease control.