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

SummaryLocal translation allows a spatial control of gene expression. Here, we performed a dual protein/mRNA localization screen, using smFISH on 523 human cell lines expressing GFP-tagged genes. A total of 32 mRNAs displayed specific cytoplasmic localizations, and we observed local translation at unexpected locations, including cytoplasmic protrusions, cell edges, endosomes, Golgi, the nuclear envelope and centrosomes, the latter being cell cycle dependent. Quantitation of mRNA distribution and automatic pattern classification revealed a high degree of localization heterogeneity between cells. Surprisingly, mRNA localization frequently required ongoing translation, indicating widespread co-translational RNA targeting. Interestingly, while P-body accumulation was frequent (15 mRNAs), four mRNAs accumulated in foci that were distinct structures. These foci lacked the mature protein, but nascent polypeptide imaging showed that they were specialized translation factories. For β-catenin, foci formation was regulated by Wnt, relied on APC-dependent polysome aggregation, and led to nascent protein degradation. Thus, translation factories uniquely regulate nascent protein metabolism and create a fine granular compartmentalization of translation.

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Subcellular mRNA localization and local translation of Arhgap11a in radial glial cells regulates cortical development

10.1101/2020.07.30.229724 ◽

2020 ◽

Cited By ~ 1

Author(s):

Louis-Jan Pilaz ◽

Kaumudi Joshi ◽

Jing Liu ◽

Yuji Tsunekawa ◽

Fernando Alsina ◽

...

Keyword(s):

Glial Cells ◽

Mrna Localization ◽

Local Translation ◽

Control Of Gene Expression ◽

Radial Glial Cells ◽

Rhoa Activity ◽

Cis Element ◽

Control Brain ◽

Radial Glial ◽

Cellular Compartments

mRNA localization and local translation enable exquisite spatial and temporal control of gene expression, particularly in highly polarized and elongated cells. These features are especially prominent in radial glial cells (RGCs), which serve as neural and glial precursors of the developing cerebral cortex, and scaffolds for migrating neurons. Yet the mechanisms by which distinct sub-cellular compartments of RGCs accomplish their diverse functions are poorly understood. Here, we demonstrate that subcellular RNA localization and translation of the RhoGAP Arhgap11a controls RGC morphology and mediates cortical cytoarchitecture. Arhgap11a mRNA and protein exhibit conserved localization to RGC basal structures in mice and humans, conferred by a 5′UTR cis-element. Proper RGC morphology relies upon active Arhgap11a mRNA transport and localization to basal structures, where ARHGAP11A is locally synthesized. Thus, RhoA activity is spatially and acutely activated via local translation in RGCs to promote neuron positioning and cortical cytoarchitecture. Altogether, our study demonstrates that mRNA localization and local translation mediate compartmentalization of neural progenitor functions to control brain development.

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A conserved choreography of mRNAs at centrosomes reveals a localization mechanism involving active polysome transport

10.1101/2020.09.04.282038 ◽

2020 ◽

Cited By ~ 1

Author(s):

Adham Safieddine ◽

Emeline Coleno ◽

Abdel-Meneem Traboulsi ◽

Oh Sung Kwon ◽

Frederic Lionneton ◽

...

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

High Throughput ◽

Local Translation ◽

Protein Coding ◽

Control Of Gene Expression ◽

Spatial Control ◽

Centrosomal Protein ◽

Protein Coding Genes ◽

Drug Treatments

AbstractLocal translation allows for a spatial control of gene expression. Here, we used high-throughput smFISH to screen centrosomal protein-coding genes, and we describe 8 human mRNAs accumulating at centrosomes. These mRNAs localize at different stages during cell cycle with a remarkable choreography, indicating a finely regulated translational program at centrosomes. Interestingly, drug treatments and reporter analyses revealed a common translation-dependent localization mechanism requiring the nascent protein. Using ASPM and NUMA1 as models, single mRNA and polysome imaging revealed active movements of endogenous polysomes towards the centrosome at the onset of mitosis, when these mRNAs start localizing. ASPM polysomes associate with microtubules and localize by either motor-driven transport or microtubule pulling. Remarkably, the Drosophila orthologs of the human centrosomal mRNAs also localize to centrosomes and also require translation. These data identify a conserved family of centrosomal mRNAs that localize by active polysomes transport mediated by nascent proteins.

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A choreography of centrosomal mRNAs reveals a conserved localization mechanism involving active polysome transport

Nature Communications ◽

10.1038/s41467-021-21585-7 ◽

2021 ◽

Vol 12 (1) ◽

Cited By ~ 2

Author(s):

Adham Safieddine ◽

Emeline Coleno ◽

Soha Salloum ◽

Arthur Imbert ◽

Abdel-Meneem Traboulsi ◽

...

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

High Throughput ◽

Local Translation ◽

Protein Coding ◽

Control Of Gene Expression ◽

Spatial Control ◽

Centrosomal Protein ◽

Protein Coding Genes ◽

Drug Treatments

AbstractLocal translation allows for a spatial control of gene expression. Here, we use high-throughput smFISH to screen centrosomal protein-coding genes, and we describe 8 human mRNAs accumulating at centrosomes. These mRNAs localize at different stages during cell cycle with a remarkable choreography, indicating a finely regulated translational program at centrosomes. Interestingly, drug treatments and reporter analyses reveal a common translation-dependent localization mechanism requiring the nascent protein. UsingASPMandNUMA1as models, single mRNA and polysome imaging reveals active movements of endogenous polysomes towards the centrosome at the onset of mitosis, when these mRNAs start localizing. ASPM polysomes associate with microtubules and localize by either motor-driven transport or microtubule pulling. Remarkably, theDrosophilaorthologs of the human centrosomal mRNAs also localize to centrosomes and also require translation. These data identify a conserved family of centrosomal mRNAs that localize by active polysome transport mediated by nascent proteins.

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mRNA Localization and Local Translation in Neurons

SSRN Electronic Journal ◽

10.2139/ssrn.3620551 ◽

2020 ◽

Author(s):

Mohammad Mofatteh

Keyword(s):

Mrna Localization ◽

Local Translation

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miranda localizes staufen and prospero asymmetrically in mitotic neuroblasts and epithelial cells in early Drosophila embryogenesis

Development ◽

10.1242/dev.125.20.4089 ◽

1998 ◽

Vol 125 (20) ◽

pp. 4089-4098 ◽

Cited By ~ 3

Author(s):

F. Matsuzaki ◽

T. Ohshiro ◽

H. Ikeshima-Kataoka ◽

H. Izumi

Keyword(s):

Epithelial Cells ◽

Rna Binding ◽

Rna Binding Protein ◽

Mrna Localization ◽

Cell Cortex ◽

Drosophila Embryogenesis ◽

Cellular Asymmetry ◽

Molecular Machinery ◽

Cycle Dependent ◽

Ganglion Mother Cell

When neuroblasts divide, prospero protein and mRNA segregate asymmetrically into the daughter neuroblast and sibling ganglion mother cell. miranda is known to localize prospero protein to the basal cell cortex of neuroblasts while the staufen RNA-binding protein mediates prospero mRNA localization. Here we show that miranda is required for asymmetric staufen localization in neuroblasts. Analyses using miranda mutants reveal that prospero and staufen interact with miranda under the same cell-cycle-dependent control. miranda thus acts to partition both prospero protein and mRNA. Furthermore, miranda localizes prospero and staufen to the basolateral cortex in dividing epithelial cells, which express the three proteins prior to neurogenesis. Our observations suggest that the epithelial cell and neuroblast (both of epithelial origin) share the same molecular machinery for creating cellular asymmetry.

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Differential regulation of mRNA fate by the human Ccr4-Not complex is driven by coding sequence composition and mRNA localization

Genome Biology ◽

10.1186/s13059-021-02494-w ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Sarah L. Gillen ◽

Chiara Giacomelli ◽

Kelly Hodge ◽

Sara Zanivan ◽

Martin Bushell ◽

...

Keyword(s):

Gene Expression ◽

Protein Synthesis ◽

Mrna Stability ◽

Mrna Localization ◽

Synthesis Rate ◽

Protein Synthesis Rate ◽

Translational Efficiency ◽

Mrna Levels ◽

Control Of Gene Expression ◽

Mrna Fate

Abstract Background Regulation of protein output at the level of translation allows for a rapid adaptation to dynamic changes to the cell’s requirements. This precise control of gene expression is achieved by complex and interlinked biochemical processes that modulate both the protein synthesis rate and stability of each individual mRNA. A major factor coordinating this regulation is the Ccr4-Not complex. Despite playing a role in most stages of the mRNA life cycle, no attempt has been made to take a global integrated view of how the Ccr4-Not complex affects gene expression. Results This study has taken a comprehensive approach to investigate post-transcriptional regulation mediated by the Ccr4-Not complex assessing steady-state mRNA levels, ribosome position, mRNA stability, and protein production transcriptome-wide. Depletion of the scaffold protein CNOT1 results in a global upregulation of mRNA stability and the preferential stabilization of mRNAs enriched for G/C-ending codons. We also uncover that mRNAs targeted to the ER for their translation have reduced translational efficiency when CNOT1 is depleted, specifically downstream of the signal sequence cleavage site. In contrast, translationally upregulated mRNAs are normally localized in p-bodies, contain disorder-promoting amino acids, and encode nuclear localized proteins. Finally, we identify ribosome pause sites that are resolved or induced by the depletion of CNOT1. Conclusions We define the key mRNA features that determine how the human Ccr4-Not complex differentially regulates mRNA fate and protein synthesis through a mechanism linked to codon composition, amino acid usage, and mRNA localization.

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CMTr cap-adjacent 2’-O-ribose mRNA methyltransferases are required for reward learning and mRNA localization to synapses

10.1101/2021.06.24.449724 ◽

2021 ◽

Author(s):

Irmgard U. Haussmann ◽

Yanying Wu ◽

Mohanakarthik P. Nallasivan ◽

Nathan Archer ◽

Zsuzsanna Bodi ◽

...

Keyword(s):

Protein Synthesis ◽

Cell Adhesion ◽

Mushroom Body ◽

Double Mutant ◽

Mrna Localization ◽

Signaling Molecules ◽

Reward Learning ◽

Local Translation ◽

Viral Mrnas ◽

Conditional Expression

AbstractCap-adjacent nucleotides of animal, protist and viral mRNAs can be dynamically O-methylated at the 2’ position of the ribose (cOMe). The functions of cOMe in animals, however, remain unknown. Here we show that the two cap methyltransferases (CMTr1 and CMTr2) of Drosophila can methylate the ribose of the first nucleotide in mRNA. Double-mutant flies lack cOMe but are viable. Consistent with prominent neuronal expression, they have a reward learning defect that can be rescued by conditional expression in mushroom body neurons before training. Among CMTr targets are cell adhesion and signaling molecules relevant for learning and cOMe is required for local translation of mRNAs at synapses. Hence, our study reveals a mechanism to co-transcriptionally prime mRNAs by cOMe for localized protein synthesis at synapses.

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Subcellular sequencing of single neurons reveals the dendritic transcriptome of GABAergic interneurons

eLife ◽

10.7554/elife.63092 ◽

2021 ◽

Vol 10 ◽

Author(s):

Julio D Perez ◽

Susanne tom Dieck ◽

Beatriz Alvarez-Castelao ◽

Georgi Tushev ◽

Ivy CW Chan ◽

...

Keyword(s):

Hippocampal Neurons ◽

Large Population ◽

Cell Types ◽

Mrna Localization ◽

Gabaergic Neurons ◽

Local Translation ◽

Single Neurons ◽

Cell Type ◽

Cell Type Specific

Although mRNAs are localized in the processes of excitatory neurons, it is still unclear whether interneurons also localize a large population of mRNAs. In addition, the variability in the localized mRNA population within and between cell-types is unknown. Here we describe the unbiased transcriptomic characterization of the subcellular compartments of hundreds of single neurons. We separately profiled the dendritic and somatic transcriptomes of individual rat hippocampal neurons and investigated mRNA abundances in the soma and dendrites of single glutamatergic and GABAergic neurons. We found that, like their excitatory counterparts, interneurons contain a rich repertoire of ~4000 mRNAs. We observed more cell type-specific features among somatic transcriptomes than their associated dendritic transcriptomes. Finally, using cell-type specific metabolic labelling of isolated neurites, we demonstrated that the processes of Glutamatergic and, notably, GABAergic neurons were capable of local translation, suggesting mRNA localization and local translation is a general property of neurons.

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