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Publications - Cell and Metabolic Biology
Publications
Plants have developed a robust transcription machinery to combat potential pathogenic organisms. One of the hallmarks of early immune responses is the activation of the WRKY transcription factors post infection. Specific WRKYs proteins from Arabidopsis are known substrates of MAPK pathway to mediate the flg22 elicited early immunity. In the current study, using the Golden Gate cloning strategy, we aim to clone the entire WRKY transcription factor family from Oryza sativa ssp. indica consisting of more than 100 members and study their MAPK interaction and subsequent role in PTI. Using a reporter LUC assay in protoplasts we investigated the early defense responses in a few interesting OsWRKY candidates. Interestingly, we observed stringent regulation of WRKY expression in cells and their transcriptional expression only under specific stress responses. The phenomenon of gene expression regulation by intron retention (IR) was prevalently observed in rice WRKY transcripts. We could show the role of WRKY8, 24, and 77 in early defense responses. It was observed that WRKY24 enhanced the expression of early defense response marker genes like NHL10 while WRKY8 and WRKY77 supressed their expression. This study highlights the complicated mechanism by which OsWRKYs expression is possibly regulated and the distinctive roles of some individual members in plant immunity. At the same time this study serves as a cautionary warning for plant researchers to be mindful of the intron retention mechanism while cloning OsWRKYs.
Publications
Type III secretion (T3S) systems are essential pathogenicity factors of most Gram-negative bacteria and translocate effector proteins into plant or animal cells. T3S systems can, therefore, be used as tools for protein delivery into eukaryotic cells, for instance after transfer of the T3S gene cluster into nonpathogenic recipient strains. Here, we report the modular cloning of the T3S gene cluster from the plant-pathogenic bacterium Xanthomonas euvesicatoria. The resulting multigene construct encoded a functional T3S system and delivered effector proteins into plant cells. The modular design of the T3S gene cluster allowed the efficient replacement and rearrangement of single genes or operons and the insertion of reporter genes for functional studies. In the present study, we used the modular T3S system to analyze the assembly of a fluorescent fusion of the predicted cytoplasmic ring protein HrcQ. Our studies demonstrate the use of the modular T3S gene cluster for functional analyses and mutant approaches in X. euvesicatoria. A potential application of the modular T3S system as protein delivery tool is discussed.
Publications
Plant Synthetic Biology requires robust and efficient methods for assembling multigene constructs. Golden Gate cloning provides a precision module-based cloning technique for facile assembly of multiple genes in one construct. We present here a versatile resource for plant biologists comprising a set of cloning vectors and 96 standardized parts to enable Golden Gate construction of multigene constructs for plant transformation. Parts include promoters, untranslated sequences, reporters, antigenic tags, localization signals, selectable markers, and terminators. The comparative performance of parts in the model plant Nicotiana benthamiana is discussed.
Publications
The induction of chitinase (CAChi2) mRNA started as early as 6 h after inoculation and gradually increased in the incompatible interaction of pepper stems with Phytophthora capsici. In the compatible interaction, the induction of the chitinase transcripts was detected later than that in the incompatible interaction. The CAChi2 mRNA was usually localized in the vascular tissues and their expression was constricted in the phloem-related cells. These results showed that the spatial pattern of CAChi2 mRNA expression was similar in both compatible and incompatible interactions but the temporal patterns were different from each other. In addition, the early induction ofCAChi2 mRNA was quite distinct in the incompatible interaction. Immunogold labelling data showed specific labelling of chitinase on the cell wall of the oomycete in both compatible and incompatible interactions at 24 h after inoculation. In particular, numerous gold particles were deposited on the cell wall of P. capsici with a predominant accumulation over areas showing signs of degradation in the incompatible interaction. Chitinase labelling was also detected in the intercellular space and the host cytoplasm. However, healthy pepper stem tissue was nearly free of labelling.
This page was last modified on 27 Jan 2025 .
