Introduction of the Ribo-BiFC method to plants using a split mVenus approach.

Abstract

Background Translation is a fundamental process for every living organism. In plants, the rate of translation is tightly modulated during development and in responses to environmental cues. However, it is challenging to measure the actual translation state of the tissues in vivo. Results Here, we report the introduction of an in vivo translation marker based on bimolecular fluorescence complementation, the Ribo-BiFC. We combined a method originally developed for the fruitflies with an improved low background split-mVenus BiFC system previously described in plants. We labelled small subunit ribosomal proteins (RPS) and large subunit ribosomal proteins (RPL) of Arabidopsis thaliana with fragments of the mVenus fluorescent protein (FP). We tested the Ribo-BiFC method using transiently expressed recombinant ribosomal proteins in epidermal cells of Nicotiana benthamiana. The BiFC-tagged ribosomal proteins complemented the mVenus molecule and were detected by fluorescence microscopy, potentially visualizing the close proximity of translating assembled 80S ribosomal subunits. Although the resulting signal is less intense than that of known interactors, its detection points to the functionality of the system. Conclusions This Ribo-BiFC approach has further potential for use in stable transgenic lines in enabling the visualisation of translational rate in plant tissues and changing translation dynamics during plant development, under abiotic stress or in different genetic backgrounds.

Code and data availability

The paper describes a Ribo-BiFC fluorescence microscopy method in plants. No public phenotype/trait datasets, plant images, author analysis code, or trained models are deposited; the authors state materials and data are available only on request, and no public repository URL is provided. Supplementary Material 1 is a D

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