ALS-associated fused in sarcoma (FUS) mutations disrupt Transportin-mediated nuclear import

Abstract

Mutations in fused in sarcoma (FUS) are a cause of familial amyotrophic lateral sclerosis (fALS). Patients carrying point mutations in the C-terminus of FUS show neuronal cytoplasmic FUS-positive inclusions, whereas in healthy controls, FUS is predominantly nuclear. Cytoplasmic FUS inclusions have also been identified in a subset of frontotemporal lobar degeneration (FTLD-FUS). We show that a non-classical PY nuclear localization signal (NLS) in the C-terminus of FUS is necessary for nuclear import. The majority of fALS-associated mutations occur within the NLS and impair nuclear import to a degree that correlates with the age of disease onset. This presents the first case of disease-causing mutations within a PY-NLS. Nuclear import of FUS is dependent on Transportin, and interference with this transport pathway leads to cytoplasmic redistribution and recruitment of FUS into stress granules. Moreover, proteins known to be stress granule markers co-deposit with inclusions in fALS and FTLD-FUS patients, implicating stress granule formation in the pathogenesis of these diseases. We propose that two pathological hits, namely nuclear import defects and cellular stress, are involved in the pathogenesis of FUS-opathies.

Astrocytes expressing ALS‐linked mutant FUS induce motor neuron death through release of tumor necrosis factor‐alpha

“…Because the redistribution of nuclear FUS has been observed in a variety of cell culture models (Bosco et al, 2010; Dormann et al, 2010; Farg et al, 2012), we examined it in our in vitro system of cultured astrocytes transduced with mutFUS. Isolated primary astrocytes were firstly tested for purity using the glutamate transporter GLAST as a marker of mature astrocytes (Supporting Information Figure S1a,c).…”

“…Over 35 mutations have been described in FUS, which has been identified as the primary cause of ALS type 6 (Vance et al, 2009). In healthy controls this multifunctional protein, with roles ranging from DNA repair, transcriptional regulation and mRNA transport (Lagier‐Tourenne, Polymenidou, & Cleveland, 2010), is located primarily in the nucleus (Dormann et al, 2010); however, post mortem tissues of FUS‐ALS patients reveal cytoplasmic inclusions in affected neurons and glia (Kwiatkowski et al, 2009). Specifically, carriers of the FUS R521C, R521G, R521H, R524W, or G507N mutations show wide‐spread FUS pathology (Blair et al, 2009; Hewitt et al, 2010; Rademakers et al, 2010), including glial and neuronal cell loss, with increasing distribution of FUS‐immunoreactive inclusions in patients with longer disease durations (Suzuki et al, 2012).…”

Linking RNA Dysfunction and Neurodegeneration in Amyotrophic Lateral Sclerosis

“…A primary abnormality in nucleocytoplasmic transport would be expected to stimulate mislocalization of TDP43 and FUS, and, as a result, disrupt RNA processing. Consistent with this hypothesis, disease-associated mutations in TARDBP and FUS promote cytoplasmic mislocalization of TDP43 and FUS, and enhance neurotoxicity [61,141]. TDP43 also regulates the expression of Ran, a master mediator of nucleocytoplasmic transport [37,142], and Ran expression was reduced to 60 % of normal levels in FTLD-TDP cortex [143].…”

“…Most ALS-associated mutations in FUS involve the nuclear localization signal [15,16], resulting in deficient nuclear import and cytoplasmic accumulation of the protein [61]. Reductions in FUS induce widespread changes in gene expression and splicing that demonstrate partial overlap with those observed in TDP43-deficient murine primary neurons and human induced pluripotent stem cell (iPSC)-derived neurons [36,59].…”

“…Cytoplasmic mislocalization of TDP43 and FUS are characteristic pathologic features in the majority of patients with ALS, and mounting evidence indicates that accumulation of either protein in the cytoplasm is neurotoxic [61,141]. Deletion of the RNA-binding domain prevents the toxicity of cytoplasmic TDP43 and FUS [39][40][41], and dbr1 deficiency creates a similar effect by flooding cells with intron lariats that compete for TDP43's RNA binding sites [114].…”