Menu

Close

A localization screen reveals translation factories and widespread co-translational RNA targeting

Racha Chouaib, Adham Safieddine, Xavier Pichon, Arthur Imbert, Oh Sung Kwon, Aubin Samacoits, Abdel-Meneem Traboulsi, Marie-Cécile Robert, Nikolay Tsanov, Emeline Coleno, Ina Poser, Christophe Zimmer, Anthony Hyman, Hervé Le Hir, Kazem Zibara, Marion Peter, Florian Mueller, Thomas Walter, Edouard Bertrand

Preprint posted on May 21, 2020 https://www.biorxiv.org/content/10.1101/2020.05.20.106989v1

mRNA localization is a heterogeneous riddle: translation might be the solution

Selected by Mafalda Pimentel

Background

Much progress has been made since the first report on localization of a specific mRNA in 1983 (non-muscle actin in the egg of the ascidian Styela1). Meanwhile, several transcripts were shown to localize in different cell types and species, usually through sequence-specific mechanisms involving RNA binding proteins and molecular motors. Some mRNA localization mechanisms are linked to translation: they can either be repressed during transport to ensure local delivery of proteins, can localize mRNAs by linkage of the ribosomes to nascent localization peptides.

Recent studies deployed “spatial transcriptome” in situ approaches and observed global patterns of distribution for various mRNAs, but lacked mechanistic insight. By screening the localization of 523 GFP-tagged transcripts, the authors of this preprint aimed to understand how local translation affects overall cellular mRNA distribution.

 

Key findings

Importantly, the use of a library of 523 HeLa cell lines containing GFP-tagged genes in bacterial artificial chromosomes (BAC)2, allowed to:

  1. use a single set of single-molecule FISH probes (GFP) to recognize the different mRNAs
  2. look at the GFP-tagged protein localization
  3. expect that all the regulatory sequences (promoter, enhancer, introns, UTRs) will faithfully reproduce the behavior of endogenous mRNAs

The authors initially screened mRNAs encoding motor proteins, and validated the observed localizations for some endogenous transcripts. They proceeded to analyze the remaining library, ending up with six patterns of localization: at the “cell edge” or specifically in “protrusions”; aggregated in cytoplasmic “foci”; in the “perinuclear” region, surrounding the “nuclear edge” or even “intranuclear”; and at the centrosome of “mitotic” cells.

 

Figure 1 – A) Schematic of the localization patterns detected in interphase cells. B)  Examples of some localized mRNAs (red) and respective GFP-tagged proteins (green). Source: figure 3 of the preprint.

 

Overall, 6% of the analyzed transcripts showed a specific cytoplasmic localization. The accumulation of mRNAs in foci was the pattern most often observed. While the majority of these mRNAs colocalized with P-bodies, the transcripts of four genes did not (BUB1, DYNC1H1, CTNNB1/β-catenin and ASPM) thus constituting distinct structures.

To perform a more quantitative and unbiased analysis, the authors adapted an mRNA detection algorithm to include mixed patterns of localization3.  Through machine learning, the localization features of 27 mRNAs in 9710 cells were thoroughly calculated. The results were similar to the manual annotations, but in addition revealed a high degree of intra and intercellular heterogeneity for all studied mRNAs.

 

Of the screened mRNAs, 2% colocalized with the GFP-tagged protein. Treatment with Puromycin (translation inhibitor that releases nascent peptide) disrupted the localization patterns for most of these mRNAs as well as the colocalization with the encoded protein. Local translation was also demonstrated (using a SunTag) for the four mRNAs accumulated in non-P-body foci, which completely disassembled when translation was inhibited and were therefore entitled as “translation factories”.

Interestingly, the few cells that did not show β-catenin translation factories, had high levels of nuclear β-catenin-GFP. Since this protein was described to enter the nucleus upon Wnt signaling, the authors stimulated cells with WNT3A and observed disassembly of translation factories. Furthermore, they showed that the degradation/oligomerization proteins Axin and APC were present in these foci and that their depletion led to its disassembly. These results suggest that the β-catenin mRNA clusters at co-translational protein degradation foci.

 

What I like about this work

First of all, I really like the experimental approach. It is known that splicing and mRNA-RBP stoichiometry can affect transcript localization. The use of genome integrated BACs allows the screening of multiple mRNA and protein localizations in conditions very similar to endogenous expression. I also appreciate the adaptation and use of an automated quantification algorithm to show the heterogeneity in mRNA distribution. This is an obvious feature that is often overlooked, although it may be important to understand how mRNA localization varies depending on the specimen and environment. This work also provides an extensive list of mRNAs that localize through translation dependent mechanisms, which can be very useful to the community.

 

Standing questions for the future

It is interesting to see that kinesin mRNAs accumulate at protrusions, particularly Kif1c, since they are plus-end motors. Could this reflect its role as an integrin recycling transporter?

Given the extended periods of ASPM polysomes at the nuclear envelope, could these represent stalled/poised translation events related to the cell cycle status?

 

References
1. Jeffery, W., Tomlinson, C. & Richard, B. Localization of actin messenger RNA during early ascidian development.Developmental Biology 99, 408–417 (1983).
2. Poser, I. et al. BAC TransgeneOmics: a high-throughput method for exploration of protein function in mammals. Nature Methods 5, 409–415 (2008).
3. Samacoits, A. et al. A computational framework to study sub-cellular RNA localization. Nature Communications 9, 4584 (2018).

Tags: foci, local translation, mrna, mrna localization, rna, smfish, translation

Posted on: 1st June 2020 , updated on: 2nd June 2020

doi: https://doi.org/10.1242/prelights.21430

Read preprint (No Ratings Yet)




  • Have your say

    Your email address will not be published. Required fields are marked *

    This site uses Akismet to reduce spam. Learn how your comment data is processed.

    Sign up to customise the site to your preferences and to receive alerts

    Register here

    Also in the cell biology category:

    Planar Cell Polarity – PCP

    This preList contains preprints about the latest findings on Planar Cell Polarity (PCP) in various model organisms at the molecular, cellular and tissue levels.

     



    List by Ana Dorrego-Rivas

    BioMalPar XVI: Biology and Pathology of the Malaria Parasite

    [under construction] Preprints presented at the (fully virtual) EMBL BioMalPar XVI, 17-18 May 2020 #emblmalaria

     



    List by Gautam Dey, Samantha Seah

    1

    Cell Polarity

    Recent research from the field of cell polarity is summarized in this list of preprints. It comprises of studies focusing on various forms of cell polarity ranging from epithelial polarity, planar cell polarity to front-to-rear polarity.

     



    List by Yamini Ravichandran

    TAGC 2020

    Preprints recently presented at the virtual Allied Genetics Conference, April 22-26, 2020. #TAGC20

     



    List by Maiko Kitaoka, Madhuja Samaddar, Miguel V. Almeida, Sejal Davla, Jennifer Ann Black, Gautam Dey

    3D Gastruloids

    A curated list of preprints related to Gastruloids (in vitro models of early development obtained by 3D aggregation of embryonic cells)

     



    List by Paul Gerald L. Sanchez and Stefano Vianello

    ECFG15 – Fungal biology

    Preprints presented at 15th European Conference on Fungal Genetics 17-20 February 2020 Rome

     



    List by Hiral Shah

    ASCB EMBO Annual Meeting 2019

    A collection of preprints presented at the 2019 ASCB EMBO Meeting in Washington, DC (December 7-11)

     



    List by Madhuja Samaddar, Ramona Jühlen, Amanda Haage, Laura McCormick, Maiko Kitaoka

    EMBL Seeing is Believing – Imaging the Molecular Processes of Life

    Preprints discussed at the 2019 edition of Seeing is Believing, at EMBL Heidelberg from the 9th-12th October 2019

     



    List by Gautam Dey

    Autophagy

    Preprints on autophagy and lysosomal degradation and its role in neurodegeneration and disease. Includes molecular mechanisms, upstream signalling and regulation as well as studies on pharmaceutical interventions to upregulate the process.

     



    List by Sandra Malmgren Hill

    Lung Disease and Regeneration

    This preprint list compiles highlights from the field of lung biology.

     



    List by Rob Hynds

    Cellular metabolism

    A curated list of preprints related to cellular metabolism at Biorxiv by Pablo Ranea Robles from the Prelights community. Special interest on lipid metabolism, peroxisomes and mitochondria.

     



    List by Pablo Ranea Robles

    BSCB/BSDB Annual Meeting 2019

    Preprints presented at the BSCB/BSDB Annual Meeting 2019

     



    List by Gautam Dey

    MitoList

    This list of preprints is focused on work expanding our knowledge on mitochondria in any organism, tissue or cell type, from the normal biology to the pathology.

     



    List by Sandra Franco Iborra

    Biophysical Society Annual Meeting 2019

    Few of the preprints that were discussed in the recent BPS annual meeting at Baltimore, USA

     



    List by Joseph Jose Thottacherry

    ASCB/EMBO Annual Meeting 2018

    This list relates to preprints that were discussed at the recent ASCB conference.

     



    List by Gautam Dey, Amanda Haage
    Close