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bioRxiv · 10.1101/706358

Visualisation of ribosomes in Drosophila axons using Ribo-BiFC

Singh, A. K.·Abdullahi, A.·Soller, M.·David, A.·Brogna, S.
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Abstract

Rates of protein synthesis and the number of translating ribosomes vary greatly between different cells in various cell states. The distribution of assembled, and potentially translating, ribosomes within cells can be visualised in Drosophila by using Bimolecular Fluorescence Complementation (BiFC) to monitor the interaction between tagged pairs of 40S and 60S ribosomal proteins (RPs) that are close neighbours across inter-subunit junctions in the assembled 80S ribosome. Here we describe transgenes that express two novel RP pairs tagged with Venus-based BiFC fragments that considerably increase the sensitivity of this technique that we termed Ribo-BiFC. This improved method should provide a convenient way of monitoring the local distribution of ribosomes in most Drosophila cells and we suggest that could be implemented in other organisms. We visualized 80S ribosomes in larval photoreceptors and in other neurons. Assembled ribosomes are most abundant in the various neuronal cell bodies, but they are also present along the lengths of axons and are concentrated in growth cones of larval and pupal photoreceptors. Surprisingly, there is relatively less puromycin incorporation in the distal portion of axons in the optic stalk, suggesting that some of the ribosomes that have started translation may not be engaged in elongation in axons that are still growing.

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Singh, A. K., Abdullahi, A., Soller, M., David, A., Brogna, S.. 2019-07-17. Visualisation of ribosomes in Drosophila axons using Ribo-BiFC. https://doi.org/10.1101/706358

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