engleski

Protein interaction analysis of BPM1, DMS3 and RDM1 in tobacco cells using bimolecular fluorescent complementation

BPM proteins are known to participate in the regulation of several transcription factors via the ubiquitin proteasome pathway. DMS3 and RDM1 proteins are important components of the RNA- directed DNA methylation (RdDM) machinery. In preliminary research, DMS3 and RDM1 were shown to be potential interactors of BPM1. Here, interactions of BPM1, DMS3 and RDM1 proteins were analyzed via bimolecular fluorescence complementation (BiFC). To perform BiFC analysis, suitable plasmid constructs were generated using In-Fusion cloning technique. Nicotiana benthamiana, Allium cepa and BY-2 cells were transiently transformed with the generated constructs via agroinfiltration or microparticle bombardment. Finally, fluorescence microscopy was used to detect and analyze protein interactions. DMS3 and RDM1 interacted in all examined cells, while DMS3 dimer formation was detected in N. benthamiana and A. cepa epidermal cells. Interaction between BPM1 and DMS3 was proved in N. benthamiana leaf epiderm and in tobacco BY-2 cells. By using BiFC, no interaction was detected between BPM1 and RDM1 in this study. Further research is required to better understand the details of interaction between BPM1 and DMS3 proteins and the potential effect of BPM1 on RdDM.

BTB ; MATH ; RdDM ; BiFC ; transformation

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