The RNA-binding protein FUS/TLS undergoes calcium-mediated nuclear egress during excitotoxic stress and is required for GRIA2 mRNA processing

Abstract

Edited by Paul E. Fraser Excitotoxic levels of glutamate represent a physiological stress that is strongly linked to amyotrophic lateral sclerosis (ALS) and other neurological disorders. Emerging evidence indicates a role for neurodegenerative disease-linked RNAbinding proteins (RBPs) in the cellular stress response. However, the relationships between excitotoxicity, RBP function, and disease have not been explored. Here, using primary cortical and motor neurons, we found that excitotoxicity induced the translocation of select ALS-linked RBPs from the nucleus to the cytoplasm within neurons. RBPs affected by excitotoxicity included TAR DNA-binding protein 43 (TDP-43) and, most robustly, fused in sarcoma/translocated in liposarcoma (FUS/ TLS or FUS). We noted that FUS is translocated through a calcium-dependent mechanism and that its translocation coincides with striking alterations in nucleocytoplasmic transport. Furthermore, glutamate-induced up-regulation of glutamate ionotropic receptor ␣-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type subunit 2 (GRIA2) in neurons depended on FUS expression, consistent with a functional role for FUS in excitotoxic stress. These findings reveal molecular links among prominent factors in neurodegenerative diseases, namely excitotoxicity, disease-associated RBPs, and nucleocytoplasmic transport. This work was supported by National Institutes of Health Grants R21 NS091860 (to D. A. B.) and R01 NS078145 (to D. A. B.) from NINDS; ALS Association Grant 18-IIA-418 (to C. F.), and a Zelda Haidek Memorial Scholarship from University of Massachusetts Medical School (to M. T.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. This article contains Figs. S1-S6.

FUS contributes to mTOR-dependent inhibition of translation

“…4 B ). Because previous reports have shown that FUS localization to the cytoplasm can be enhanced in response to translation repression ( 36 , 42 , 70 ), we examined the cytoplasmic distribution of FUS in response to Torin1. Using subcellular fractionation of control or Torin1-treated cells, we observed a modest but statistically significant increase in FUS in the cytoplasmic fraction ( Fig.…”

“…ALS-linked FUS mutations in the proline-tyrosine nuclear localization sequence domain localize predominantly to the cytoplasm, which correlate with a reduction in protein synthesis ( 20 , 33 , 34 ). Changes in mRNA trafficking and stability ( 23 , 24 , 35 , 36 ), as well as aberrant protein–protein interactions ( 37 , 38 , 39 ), have all been attributed to these pathological FUS mutations. Although WT FUS is mainly localized in the nucleus, its localization to the cytoplasm can also be enhanced in response to conditions that restrict translation.…”

“…Although WT FUS is mainly localized in the nucleus, its localization to the cytoplasm can also be enhanced in response to conditions that restrict translation. In neurons, glutamate excitotoxicity has been shown to induce the cytoplasmic localization of FUS bound to glutamate ionotropic receptor AMPA type subunit 2 (GRIA2) mRNA, which corresponds to reduced global protein synthesis ( 36 ). In other conditions where protein synthesis is repressed, such as heat shock, sodium arsenite, or sorbitol treatments, FUS is found in cytoplasmic stress granules composed of messenger ribonucleoproteins and stalled mRNAs ( 40 , 41 , 42 , 43 ).…”

Interplay of RNA-Binding Proteins and microRNAs in Neurodegenerative Diseases

“…Dysregulation of the miR-17-92 cluster has been shown to contribute to the particular susceptibility of motor neurons in ALS model mice [ 133 ]. The cytoplasmic expression of hnRNP A1 has been reported to be increased in response to excitotoxic stress in primary cortical neurons, to a lesser extent compared to the FUS effect, and the cytoplasmic expression of hnRNP A1 may be linked to ALS and FTD by promoting protein aggregation [ 134 ]. Interestingly, the expression levels of hnRNP A1 and its regulatory factor miR-590-3p have been reported to be altered in blood cells from patients with FTLD [ 135 ].…”

Phenotypic Suppression of ALS/FTD-Associated Neurodegeneration Highlights Mechanisms of Dysfunction

“…Whether granules formed by protein overexpression are bona fide SGs is unclear, as these granules exhibit different dynamic properties compared with SGs formed in re- sponse to acute stress. Furthermore, disease-relevant stressors, such as glutamate-induced excitotoxicity, did not induce SG assembly in neurons; thus, not all types of stress necessarily result in SG formation (Tischbein et al, 2019). A recent study in Drosophila provided novel proof of concept that physical trauma can serve as an acute form of stress that triggers SG assembly in vivo (Anderson et al, 2018).…”