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ALS-linked cytoplasmic FUS assemblies are compositionally different from physiological stress granules and sequester hnRNPA3, a novel modifier of FUS toxicity

An, Haiyan; Litscher, Gioana; Watanabe, Naruaki; Wei, Wenbin; Hashimoto, Tadafumi; Iwatsubo, Takeshi; Buchman, Vladimir L.; Shelkovnikova, Tatyana A.

ALS-linked cytoplasmic FUS assemblies are compositionally different from physiological stress granules and sequester hnRNPA3, a novel modifier of FUS toxicity Thumbnail


Authors

Haiyan An

Gioana Litscher

Naruaki Watanabe

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Dr Wenbin Wei wenbin.wei2@durham.ac.uk
Chief Experimental Officer (Bioinformatics)

Tadafumi Hashimoto

Takeshi Iwatsubo

Vladimir L. Buchman

Tatyana A. Shelkovnikova



Abstract

Formation of cytoplasmic RNA-protein structures called stress granules (SGs) is a highly conserved cellular response to stress. Abnormal metabolism of SGs may contribute to the pathogenesis of (neuro)degenerative diseases such as amyotrophic lateral sclerosis (ALS). Many SG proteins are affected by mutations causative of these conditions, including fused in sarcoma (FUS). Mutant FUS variants have high affinity to SGs and also spontaneously form de novo cytoplasmic RNA granules. Mutant FUS-containing assemblies (mFAs), often called “pathological SGs”, are proposed to play a role in ALS-FUS pathogenesis. However, structural differences between mFAs and physiological SGs remain largely unknown therefore it is unclear whether mFAs can functionally substitute for SGs and how they affect cellular stress responses. Here we used affinity purification to isolate mFAs and physiological SGs and compare their protein composition. We found that proteins within mFAs form significantly more physical interactions than those in SGs however mFAs fail to recruit many factors involved in signal transduction. Furthermore, we found that proteasome subunits and certain nucleocytoplasmic transport factors are depleted from mFAs, whereas translation elongation, mRNA surveillance and splicing factors as well as mitochondrial proteins are enriched in mFAs, as compared to SGs. Validation experiments for a mFA-specific protein, hnRNPA3, confirmed its RNA-dependent interaction with FUS and its sequestration into FUS inclusions in cultured cells and in a FUS transgenic mouse model. Silencing of the Drosophila hnRNPA3 ortholog was deleterious and potentiated human FUS toxicity in the retina of transgenic flies. In conclusion, we show that SG-like structures formed by mutant FUS are structurally distinct from SGs, prone to persistence, likely cannot functionally replace SGs, and affect a spectrum of cellular pathways in stressed cells. Results of our study support a pathogenic role for cytoplasmic FUS assemblies in ALS-FUS.

Citation

An, H., Litscher, G., Watanabe, N., Wei, W., Hashimoto, T., Iwatsubo, T., …Shelkovnikova, T. A. (2022). ALS-linked cytoplasmic FUS assemblies are compositionally different from physiological stress granules and sequester hnRNPA3, a novel modifier of FUS toxicity. Neurobiology of Disease, 162, Article 105585. https://doi.org/10.1016/j.nbd.2021.105585

Journal Article Type Article
Acceptance Date Dec 11, 2021
Online Publication Date Dec 14, 2021
Publication Date 2022-01
Deposit Date Feb 16, 2022
Publicly Available Date Feb 16, 2022
Journal Neurobiology of Disease
Print ISSN 0969-9961
Publisher Elsevier
Peer Reviewed Peer Reviewed
Volume 162
Article Number 105585
DOI https://doi.org/10.1016/j.nbd.2021.105585

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