BackgroundMutations in calcium-responsive transactivator (CREST) encoding gene have been recently linked to ALS. Similar to several proteins implicated in ALS, CREST contains a prion-like domain and was reported to be a component of paraspeckles.ResultsWe demonstrate that CREST is prone to aggregation and co-aggregates with FUS but not with other two ALS-linked proteins, TDP-43 and TAF15, in cultured cells. Aggregation of CREST affects paraspeckle integrity, probably by trapping other paraspeckle proteins within aggregates. Like several other ALS-associated proteins, CREST is recruited to induced stress granules. Neither of the CREST mutations described in ALS alters its subcellular localization, stress granule recruitment or detergent solubility; however Q388stop mutation results in elevated steady-state levels and more frequent nuclear aggregation of the protein. Both wild-type protein and its mutants negatively affect neurite network complexity of unstimulated cultured neurons when overexpressed, with Q388stop mutation being the most deleterious. When overexpressed in the fly eye, wild-type CREST or its mutants lead to severe retinal degeneration without obvious differences between the variants.ConclusionsOur data indicate that CREST and certain other ALS-linked proteins share several features implicated in ALS pathogenesis, namely the ability to aggregate, be recruited to stress granules and alter paraspeckle integrity. A change in CREST levels in neurons which might occur under pathological conditions would have a profound negative effect on neuronal homeostasis.Electronic supplementary materialThe online version of this article (doi:10.1186/s13024-015-0014-y) contains supplementary material, which is available to authorized users.

Aggregation of fused in sarcoma (FUS) protein, and mutations in FUS gene, are causative to a range of neurodegenerative disorders including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia. To gain insights into the molecular mechanism whereby FUS causes neurodegeneration, we generated transgenic Drosophila melanogaster overexpressing human FUS in the photoreceptor neurons, which exhibited mild retinal degeneration. Expression of familial ALS-mutant FUS aggravated the degeneration, which was associated with an increase in cytoplasmic localization of FUS. A carboxy-terminally truncated R495X mutant FUS also was localized in cytoplasm, whereas the degenerative phenotype was diminished. Double expression of R495X and wild-type FUS dramatically exacerbated degeneration, sequestrating wild-type FUS into cytoplasmic aggregates. Notably, replacement of all tyrosine residues within the low-complexity domain, which abolished self-assembly of FUS, completely eliminated the degenerative phenotypes. Taken together, we propose that self-assembly of FUS through its low-complexity domain contributes to FUS-induced neurodegeneration.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by progressive and selective loss of motor neurons. Causative genes for familial ALS (fALS), e.g. TARDBP or FUS/TLS, have been found, among which mutations within the profilin 1 (PFN1) gene have recently been identified in ALS18. To elucidate the mechanism whereby PFN1 mutations lead to neuronal death, we generated transgenic Drosophila melanogaster overexpressing human PFN1 in the retinal photoreceptor neurons. Overexpression of wild-type or fALS mutant PFN1 caused no degenerative phenotypes in the retina. Double overexpression of fALS mutant PFN1 and human TDP-43 markedly exacerbated the TDP-43-induced retinal degeneration, i.e. vacuolation and thinning of the retina, whereas co-expression of wild-type PFN1 did not aggravate the degenerative phenotype. Notably, co-expression of TDP-43 with fALS mutant PFN1 increased the cytoplasmic localization of TDP-43, the latter remaining in nuclei upon co-expression with wild-type PFN1, whereas co-expression of TDP-43 lacking the nuclear localization signal with the fALS mutant PFN1 did not aggravate the retinal degeneration. Knockdown of endogenous Drosophila PFN1 did not alter the degenerative phenotypes of the retina in flies overexpressing wild-type TDP-43. These data suggest that ALS-linked PFN1 mutations exacerbate TDP-43-induced neurodegeneration in a gain-of-function manner, possibly by shifting the localization of TDP-43 from nuclei to cytoplasm. Amyotrophic lateral sclerosis (ALS)2 is a progressive neurodegenerative disorder characterized by selective loss of upper and lower motor neurons. Approximately 10% of ALS cases are inherited as an autosomal dominant trait (familial ALS; fALS), and mutations in a number of causative genes, e.g. Cu/Zn superoxide dismutase (1) and TARDBP (2-5), have been identified in fALS cases. Recently mutations within the PFN1 gene were identified in pedigrees of ALS 18 (OMIM614808) (6, 7). Moreover, the E117G variant in PFN1 is also considered a genetic risk factor for ALS (8, 9). However, the mechanisms whereby PFN1 mutations induce degeneration and death of motor neurons remain elusive. PFN1 is a protein implicated in the regulation of actin assembly by binding to monomeric actin (10). Homozygous PFN1 knock-out mice die shortly after fertilization (11), and brain-specific conditional knock-out of the PFN1 gene during development led to cerebellar hypoplasia and abnormal neuronal migration (12), indicating the roles of PFN1 in development. However, there have been few reports on the pathological functions of fALS mutant PFN1; the level of PFN1-bound actin was shown to be reduced upon overexpression of the fALS mutant PFN1 compared with wild-type (wt) PFN1 in HEK293 cells (6). A recent report (13) showed that the fALS mutant PFN1 failed to restore growth of PFN1 mutant yeast. These data suggested a partial loss-of-function mechanism in neurodegeneration caused by fALS mutations in PFN1.TAR DNA-binding protein of 43 kDa (TDP-43) is a member of hetero...

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