FUS Phase Separation Is Modulated by a Molecular Chaperone and Methylation of Arginine Cation-π Interactions

Abstract

SummaryReversible phase separation underpins the role of FUS in ribonucleoprotein granules and other membrane-free organelles and is, in part, driven by the intrinsically disordered low-complexity (LC) domain of FUS. Here, we report that cooperative cation-π interactions between tyrosines in the LC domain and arginines in structured C-terminal domains also contribute to phase separation. These interactions are modulated by post-translational arginine methylation, wherein arginine hypomethylation strongly promotes phase separation and gelation. Indeed, significant hypomethylation, which occurs in FUS-associated frontotemporal lobar degeneration (FTLD), induces FUS condensation into stable intermolecular β-sheet-rich hydrogels that disrupt RNP granule function and impair new protein synthesis in neuron terminals. We show that transportin acts as a physiological molecular chaperone of FUS in neuron terminals, reducing phase separation and gelation of methylated and hypomethylated FUS and rescuing protein synthesis. These results demonstrate how FUS condensation is physiologically regulated and how perturbations in these mechanisms can lead to disease.

RNAs, Phase Separation, and Membrane‐Less Organelles: Are Post‐Transcriptional Modifications Modulating Organelle Dynamics?

“…In support of the notion that local concentrations of negative charge play major roles for LLPS, nucleic acid‐mimicking biopolymers such as poly‐ADP‐ribose (PAR) induced phase transitions in vitro and in vivo and phosphorylation of low complexity regions in aggregation‐prone proteins disrupted their phase separation . Furthermore, also post‐translational modifications (i.e., methylation) affecting the charge of particular amino acids in IDR‐containing proteins disrupted cation‐pi interactions thereby modulating LLPS . Moreover, introducing additional negative charge to PLD‐containing proteins enhanced phase separation mediated by intermolecular interactions among PLDs and RRMs .…”

“…Intriguingly, while the contribution of the biophysical characteristics of RNA to these processes is being increasingly appreciated, it remains unclear how particular RNA properties are modulated at the level of individual RNAs. In the following paragraphs, we list the known characteristics and effects of post‐transcriptional chemical modifications in RNA and discuss how such modifications could be a means to dynamically regulate RNA properties during LLPS and in analogy to the recently reported roles for post‐translational modifications of proteins in the process …”

“…In addition, transcriptome‐wide mapping of in vivo RNA structures revealed that m 6 A was indeed enriched in unstructured RNA regions, indicating the potential for particular RNA modifications in regulating structure. Of note, m 5 C changes the dipole moment of the modified base, thereby impacting the aromatic amino acid‐cation interactions, which have been shown to influence LLPS . Furthermore, particular methyl groups (i.e., m 5 C or m 6 A) introduce a hydrophobic moiety to the RNA major groove, which cannot be fully solvated.…”

Biomolecular condensates in neurodegeneration and cancer

“…The interaction between FUS and its import receptor, karyopherin‐ß2 (Karβ2)/importin‐ß2, significantly diminishes FUS aggregation in vitro . The proline/tyrosine‐nuclear localization signal of FUS is frequently mutated in ALS, precluding the FUS/Karβ2 interaction and accelerating FUS aggregation (Figure C). Inhibition of the molecular chaperone, HSP70, also slows the internal dynamics of cellular SGs .…”

“…Inhibition of the molecular chaperone, HSP70, also slows the internal dynamics of cellular SGs . Interestingly, chaperone inhibition also delays SG disassembly post‐stress, suggesting that SG persistence and aggregation may be linked. Supporting this, constitutively inducing SG protein phase separation in cells (using optogenetic oligomerization tags) causes their aggregation .…”