High-throughput identification of RNA localization elements in neuronal cells

Arora, Ankita; Castro-Gutiérrez, Roberto; Moffatt, Charlie; Eletto, Davide; Becker, Raquel; Brown, Maya; Moor, Andreas E.; Russ, Holger A.; Taliaferro, J. Matthew

doi:10.1093/nar/gkac763

“…Even more overlap was observed between our subcellular proteome and two independent synapse transcriptomes ( Fig 2D ) [36, 37]. This intersection suggests that there may be a cohort of conserved genes required for maintenance of distal, subcellular structures [38, 39]. Furthermore, the non-overlapping genes may provide insights into RGC specific components and functions.…”

Section: Resultsmentioning

confidence: 96%

Subcellular proteome of radial glia reveals non-muscle myosins control basal endfeet to mediate interneuron organization

D’Arcy

¹

,

Lennox

²

,

Musso

³

et al. 2022

Preprint

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Radial glial cells (RGCs) are essential for the generation and organization of neurons in the cerebral cortex. RGCs have an elongated bipolar morphology with basal and apical endfeet which reside in distinct niches. Yet, how this subcellular compartmentalization of RGCs controls cortical development is largely unknown. Here, we employ in vivo proximity labeling using unfused BirA to generate the first subcellular proteome of RGCs and uncover new principles governing local control of cortical development. We discover a cohort of proteins that are significantly enriched in RGC basal endfeet, with MYH9 and MYH10 among the most abundant. Myh9 and Myh10 transcripts also localize to endfeet with distinct temporal dynamics. Although they each encode isoforms of nonmuscle myosin II heavy chain, Myh9 and Myh10 have drastically different requirements for RGC integrity. Myh9 loss from RGCs decreases branching complexity and causes endfoot protrusion through the basement membrane. In contrast, Myh10 controls endfoot adhesion, as mutants have unattached apical and basal endfeet. Finally, we show that Myh9- and Myh10-mediated regulation of RGC complexity and endfoot position non-cell autonomously controls interneuron number and organization in the marginal zone. Our study demonstrates the utility of in vivo proximity labeling for dissecting local control of complex systems, and reveals new mechanisms for dictating RGC integrity and cortical architecture.

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“…It has been previously shown that GA-rich motifs mediate targeting of Rab13 mRNA targeting to the actin rich cellular protrusions 37, 38 . Similarly, GA-rich motifs in Net1 mRNA were shown to mediate Net1 targeting to neurites 39 . Our RNAseq analysis of purified MBs showed Rab13 and Net1 mRNAs are also enriched in the MB, suggesting that the GA-rich motifs may also mediate CHMP4B mRNA targeting to the MBs.…”

Section: Discussionmentioning

confidence: 94%

The Novel Role of Midbody-Associated mRNAs in Regulating Abscission

Farmer

¹

,

Vaeth

²

,

Han

³

et al. 2022

Preprint

Self Cite

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Midbodies (MBs) have been shown to function during telophase as a recruiting hub, especially for ESCRT-III complex subunits, including CHMP4B, to regulate the abscission step of cytokinesis. However, the molecular machinery governing specific protein targeting and activation at the MB remains poorly understood. Until recently, it was thought that abscission regulating proteins, such as ESCRT-III complex subunits, accumulate at the MB by directly or indirectly binding to the MB resident protein, CEP55. However, recent studies have shown that depletion of CEP55 does not fully block ESCRT-III targeting to the MB, and cells in CEP55 knock-out mice divide normally. Additionally, since MBs are microtubule-rich, proteinaceous structures, it is conceptually hard to imagine how large protein complexes, such as the ESCRT-III complex, can successfully diffuse into the MB from the cytosol in a rapid and highly regulated manner. Here, we show that MBs contain mRNAs and that these MB-associated mRNAs can be locally translated, thus, resulting in the accumulation of abscission-regulating proteins. We also demonstrate that localized MB-associated translation of CHMP4B is required for it’s targeting to the abscission site. Finally, we demonstrate that 3’-UTR-dependent CHMP4B mRNA targeting to the MB is required for successful completion of cytokinesis. Based on all this data, we propose a novel method of regulating cytokinesis and abscission by MB-associated targeting and localized translation of selective mRNAs.

show abstract

“…It has been previously shown that GA-rich motifs mediate targeting of Rab13 mRNA targeting to the actin rich cellular protrusions 37,38 . Similarly, GA-rich motifs in Net1 mRNA were shown to mediate Net1 targeting to neurites 39 . Our RNAseq analysis of puri ed MBs showed Rab13 and Net1 mRNAs are also enriched in the MB, suggesting that the GA-rich motifs may also mediate CHMP4B mRNA targeting to the MBs.…”

Section: Discussionmentioning

confidence: 94%

The Novel Role of Midbody-Associated mRNAs in Regulating Abscission

Farmer

¹

,

Han

²

,

Vaeth

³

et al. 2022

Preprint

0

View full text Add to dashboard Cite

Midbodies (MBs) have been shown to function during telophase as a recruiting hub, especially for ESCRT-III complex subunits, including CHMP4B, to regulate the abscission step of cytokinesis. However, the molecular machinery governing specific protein targeting and activation at the MB remains poorly understood. Until recently, it was thought that abscission regulating proteins, such as ESCRT-III complex subunits, accumulate at the MB by directly or indirectly binding to the MB resident protein, CEP55. However, recent studies have shown that depletion of CEP55 does not fully block ESCRT-III targeting to the MB, and cells in CEP55 knock-out mice divide normally. Additionally, since MBs are microtubule-rich, proteinaceous structures, it is conceptually hard to imagine how large protein complexes, such as the ESCRT-III complex, can successfully diffuse into the MB from the cytosol in a rapid and highly regulated manner. Here, we show that MBs contain mRNAs and that these MB-associated mRNAs can be locally translated, thus, resulting in the accumulation of abscission-regulating proteins. We also demonstrate that localized MB-associated translation of CHMP4B is required for it’s targeting to the abscission site. Finally, we demonstrate that 3’-UTR-dependent CHMP4B mRNA targeting to the MB is required for successful completion of cytokinesis. Based on all this data, we propose a novel method of regulating cytokinesis and abscission by MB-associated targeting and localized translation of selective mRNAs.

show abstract

High-throughput identification of RNA localization elements in neuronal cells

Cited by 26 publications

References 50 publications

Subcellular proteome of radial glia reveals non-muscle myosins control basal endfeet to mediate interneuron organization

Subcellular proteome of radial glia reveals non-muscle myosins control basal endfeet to mediate interneuron organization

The Novel Role of Midbody-Associated mRNAs in Regulating Abscission

The Novel Role of Midbody-Associated mRNAs in Regulating Abscission

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