Abstract
Fused in sarcoma/translocated in liposarcoma (FUS/TLS) is one of causative genes for familial amyotrophic lateral sclerosis (ALS). In order to identify binding partners for FUS/TLS, we performed a yeast two-hybrid screening and found that protein arginine methyltransferase 1 (PRMT1) is one of binding partners primarily in the nucleus. In vitro and in vivo methylation assays showed that FUS/TLS could be methylated by PRMT1. The modulation of arginine methylation levels by a general methyltransferase inhibitor or conditional over-expression of PRMT1 altered slightly the nucleus-cytoplasmic ratio of FUS/TLS in cell fractionation assays. Although co-localized primarily in the nucleus in normal condition, FUS/TLS and PRMT1 were partially recruited to the cytoplasmic granules under oxidative stress, which were merged with stress granules (SGs) markers in SH-SY5Y cell. C-terminal truncated form of FUS/TLS (FUS-dC), which lacks C-terminal nuclear localization signal (NLS), formed cytoplasmic inclusions like ALS-linked FUS mutants and was partially co-localized with PRMT1. Furthermore, conditional over-expression of PRMT1 reduced the FUS-dC-mediated SGs formation and the detergent-insoluble aggregates in HEK293 cells. These findings indicate that PRMT1-mediated arginine methylation could be implicated in the nucleus-cytoplasmic shuttling of FUS/TLS and in the SGs formation and the detergent-insoluble inclusions of ALS-linked FUS/TLS mutants.
Highlights
Amyotrophic Lateral Sclerosis (ALS) is the most frequent form of adult-onset fatal progressive motor neuron diseases, characterized by the degeneration of upper and lower motor neurons
Since protein arginine methyltransferase 1 (PRMT1) is a major arginine methyltransferase modulating the functions of substrates through methylation [21], we examine whether fused in sarcoma/translocated in liposarcoma (FUS/TLS) is methylated by PRMT1 using antiDimethyl Arginine (DMA) antibody or Anti-dimethyl-Arginine
We examined whether endogenous FUS/TLS and PRMT1 are co-localized in the cytoplasmic inclusions under stress condition using SH-SY5Y cells
Summary
Amyotrophic Lateral Sclerosis (ALS) is the most frequent form of adult-onset fatal progressive motor neuron diseases, characterized by the degeneration of upper and lower motor neurons. Postmortem analysis in ALS patients showed abnormal cytoplasmic aggregations of these DNA/RNA binding proteins in affected neurons [1,2,3,4,6]. These results indicate that gain of toxic function and/ or loss of function of these DNA/RNA binding proteins might be implicated in the pathogenesis of ALS [6,7,8]. Mutations in TDP-43 and FUS/TLS gene were identified in the cases of frontotemporal lobar degeneration (FLTD), and ubiquitinpositive inclusions in some cases of FTLD contain these protein products as a major component [2,9,10,11,12], supporting that ALS and FTLD might be a part of a clinical, pathologic, and genetic disease spectrum
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