Monomethylated and unmethylated FUS exhibit increased binding to Transportin and distinguish FTLD-FUS from ALS-FUS

Abstract

Deposition of the nuclear DNA/RNA-binding protein Fused in sarcoma (FUS) in cytosolic inclusions is a common hallmark of some cases of frontotemporal lobar degeneration (FTLD-FUS) and amyotrophic lateral sclerosis (ALS-FUS). Whether both diseases also share common pathological mechanisms is currently unclear. Based on our previous finding that FUS deposits are hypomethylated in FTLD-FUS but not in ALS-FUS, we have now investigated whether genetic or pharmacological inactivation of Protein arginine methyltransferase 1 (PRMT1) activity results in unmethylated FUS or in alternatively methylated forms of FUS. To do so, we generated FUS-specific monoclonal antibodies that specifically recognize unmethylated arginine (UMA), monomethylated arginine (MMA) or asymmetrically dimethylated arginine (ADMA). Loss of PRMT1 indeed not only results in an increase of UMA FUS and a decrease of ADMA FUS, but also in a significant increase of MMA FUS. Compared to ADMA FUS, UMA and MMA FUS exhibit much higher binding affinities to Transportin-1, the nuclear import receptor of FUS, as measured by pull-down assays and isothermal titration calorimetry. Moreover, we show that MMA FUS occurs exclusively in FTLD-FUS, but not in ALS-FUS. Our findings therefore provide additional evidence that FTLD-FUS and ALS-FUS are caused by distinct disease mechanisms although both share FUS deposits as a common denominator.

Protein Arginine Methyltransferase Product Specificity Is Mediated by Distinct Active-site Architectures

“…For example, whether a particular arginine residue on histone tails is asymmetrically or symmetrically dimethylated can lead to gene repression or activation (13)(14)(15)(16)(17). However, few studies have been conducted to determine the role of MMA marks (18). It has been proposed that MMA marks are used mainly as precursors for dimethylation by the various type I and II PRMTs (17,19).…”

Aggregation of FET Proteins as a Pathological Change in Amyotrophic Lateral Sclerosis

“…In ALS-FUS but not FTLD-FUS cases, the pathological inclusions contained FUS with asymmetrically dimethylated arginines (ADMA) (Dormann et al 2012). In contrast, unmethylated/monomethylated Arg (UMA/MMA) at RGG3 region of FUS constituted the inclusions observed in FTLD-FUS but not in ALS-FUS cases (Suarez-Calvet et al 2016). Methylations of Arg residues in FUS have been shown to reduce its binding to transportin and thus suppress the nuclear import of FUS (Dormann et al 2012;Suarez-Calvet et al 2016).…”

“…While the regulation of the nuclearcytoplasmic shuttling of the FET proteins remains obscure, the nuclear import of the FET proteins is negatively regulated by chemical modifications near the PY-NLS, such as the methylation of Arg (Dormann et al 2012;Belyanskaya et al 2003) and the phosphorylation of Tyr (Leemann-Zakaryan et al 2011). Given that cytoplasmic accumulation of FUS is observed in the ALS-FUS cases, post-translational modifications affecting the function of PY-NLS may also have pathological roles in ALS (Dormann et al 2012;Suarez-Calvet et al 2016). Also, notably, significant concentrations of FUS are observed in the dendritic spines of neurons, where mRNAs are stored for their local translation upon synaptic activity (Fujii et al 2005).…”

Verbesserung der Dispersion der chemischen Verschiebungen von unstrukturierten Proteinen durch einen kovalent gebundenen Lanthanoidkomplex

“…Dabei wurde das 110 kDa große Protein Tr ansportin-1 hinzutitriert, vom dem gezeigt wurde,d ass es den C-Terminus von FUS bindet, [22] aber neueste Erkenntnisse weisen auch auf eine Rolle der RGGRegion bei der Bindung hin. [18,23] …”