Sporadic ALS has compartment-specific aberrant exon splicing and altered cell–matrix adhesion biology

Rabin, Stuart J.; Kim, Jae Mun â€ ̃Hugoâ€TM; Baughn, Michael; Libby, Ryan T.; Kim, Young Joo; Fan, Yuxin; Libby, Randell T.; Spada, Albert La; Stone, Brad; Ravits, John

doi:10.1093/hmg/ddp498

Cited by 123 publications

(126 citation statements)

References 76 publications

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“…The identification of these potential FUS-R521C target genes provides important insights into the dendritic and synaptic phenotypes in FUS-R521C mice and explain why BDNF can only partially restore the dendritic phenotype in neurons expressing FUS-R521C (Figure 8). The RNA-seq approach also reveals many target genes in the extracellular matrix assembly GO categories (GO:0005581, GO:0005201, GO:0005578, and GO:0031012) that have also been shown to be transcriptional targets of DNA damage response genes CSB and HDAC1 (43) and are frequently misregulated and misspliced in the motor neurons of SALS patients (54). Although these results are correlative, they raise the interesting possibility that the recruitment of FUS, HDAC1, and CSB may constitute a critical step in the repair of damaged DNA in FALS caused by FUS mutations and in SALS.…”

Section: Methodsmentioning

confidence: 99%

ALS-associated mutation FUS-R521C causes DNA damage and RNA splicing defects

et al. 2014

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Autosomal dominant mutations of the RNA/DNA binding protein FUS are linked to familial amyotrophic lateral sclerosis (FALS); however, it is not clear how FUS mutations cause neurodegeneration. Using transgenic mice expressing a common FALS-associated FUS mutation (FUS-R521C mice), we found that mutant FUS proteins formed a stable complex with WT FUS proteins and interfered with the normal interactions between FUS and histone deacetylase 1 (HDAC1). Consequently, FUS-R521C mice exhibited evidence of DNA damage as well as profound dendritic and synaptic phenotypes in brain and spinal cord. To provide insights into these defects, we screened neural genes for nucleotide oxidation and identified brain-derived neurotrophic factor (Bdnf ) as a target of FUS-R521C-associated DNA damage and RNA splicing defects in mice. Compared with WT FUS, mutant FUS-R521C proteins formed a more stable complex with Bdnf RNA in electrophoretic mobility shift assays. Stabilization of the FUS/Bdnf RNA complex contributed to Bdnf splicing defects and impaired BDNF signaling through receptor TrkB. Exogenous BDNF only partially restored dendrite phenotype in FUS-R521C neurons, suggesting that BDNF-independent mechanisms may contribute to the defects in these neurons. Indeed, RNA-seq analyses of FUS-R521C spinal cords revealed additional transcription and splicing defects in genes that regulate dendritic growth and synaptic functions. Together, our results provide insight into how gain-of-function FUS mutations affect critical neuronal functions. IntroductionAutosomal dominant mutations in RNA/DNA binding protein fused in sarcoma/translocated in liposarcoma (FUS/TLS) have been causally linked to familial ALS (FALS). The main pathological features in FALS with FUS mutations are FUS-positive protein aggregates in neuronal cytoplasm and dendrites (1, 2). While the majority of these aggregates are identified in the spinal motor neurons, FUS-positive aggregates have also been found in neurons in cerebral cortex and brainstem nuclei (3, 4), raising the possibility that FUS mutations may have a broader impact on the functions of other neuronal subtypes. Consistent with these observations, a subset of FALS-FUS patients also exhibits cognitive impairments during the developmental or degenerative processes.Although the exact mechanism of FUS mutations remains unclear, several previous studies have provided compelling evidence that FUS can regulate DNA damage response, transcription, and RNA processing. For instance, fibroblasts and lymphocytes from fus-null mice show increased sensitivity to ionizing irradiation and genomic instability, respectively (5, 6). Consistent with these results, our recent study shows that WT FUS proteins are rapidly recruited to DNA damage foci in neurons, where it interacts with histone deacetylase 1 (HDAC1), a critical component in

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Section: Methodsmentioning

confidence: 99%

ALS-associated mutation FUS-R521C causes DNA damage and RNA splicing defects

et al. 2014

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“…However, aberrant intron inclusion has not been reported in fALS patients. Interestingly, a study used exon arrays to probe changes in sporadic ALS patients and found aberrant splicing in cell adhesion and cell matrix genes (Rabin et al 2010). However, the exact nature of the missplicing was not determined, nor is it known whether Fus was mutated in the patients.…”

Section: Discussionmentioning

confidence: 99%

fus/TLS orchestrates splicing of developmental regulators during gastrulation

Dichmann

Harland²

2012

Genes Dev.

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Here we investigated the function of the atypical RNA-binding protein fus/TLS (fused in sarcoma/translocated in sarcoma) during early frog development. We found that fus is necessary for proper mRNA splicing of a set of developmental regulatory genes during early frog development and gastrulation. Upon fus knockdown, embryos fail to gastrulate and show mesodermal differentiation defects that we connect to intron retention in fgf8 (fibroblast growth factor 8) and fgfr2 (fgf receptor 2) transcripts. During gastrulation, the animal and marginal regions dissociate, and we show that this is caused, at least in part, by intron retention in cdh1 transcripts. We confirm the specificity of splicing defects at a genomic level using analysis of RNA sequencing (RNA-seq) and show that 3%-5% of all transcripts display intron retention throughout the pre-mRNA. By analyzing gene ontology slim annotations, we show that the affected genes are enriched for developmental regulators and therefore represent a biologically coherent set of targets for fus regulation in embryogenesis. This shows that fus is central to embryogenesis and may provide information on its function in neurodegenerative disease.[Keywords: Xenopus; fus; splicing; signaling; embryo development; RNA-seq] Supplemental material is available for this article. Pre-mRNA splicing, alternative and constitutive, is catalyzed by the spliceosome that contains five small nuclear ribonucleoproteins (snRNPs): U1, U2, U4, U5, and U6. Splicing occurs in a stepwise fashion, and a series of distinct complexes form in the process at the 59 and 39 splice sites and branch point site (Wahl et al. 2009). However, these sequences are short and poorly conserved. During splicing, the spliceosome must therefore succeed in recognizing and joining splice sites that are surrounded by numerous similar sequences and often separated by considerable distances. Further complicating the task, the interactions between the snRNPs and the pre-mRNA are generally weak and rely for specificity on numerous auxiliary proteins that interact with the core snRNPs. This general principle of multiple interactions is important not only for precisely finding correct splice pairs, but also for the flexibility necessary for alternative splice site selection.Comprehensive analysis of purified early spliceosomal complexes shows that they consist of at least 85 and perhaps as many as 150-300 different proteins with a combined mass of 2.7 MDa. Many of these function in the enzymatic and conformational changes that occur during splicing, while others are involved in exon and intron definition by binding to splicing enhancers or silencers present in the pre-mRNA. For example, members of the serine-arginine-rich splice factors (SR proteins) frequently bind sequences in exons and stimulate exon inclusion by stabilizing U1 and U2 binding to the 59 splice site and branch point site, respectively. Other well-known splicing regulators are the heterologous nuclear RNPs (hnRNPs) that generally repress exon inclusion and hence co...

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“…In addition, RNA profiling suggested direct induction of C1q complement components in laser-microdissected postmortem ALS motor neurons (26). With respect to ALS mouse models, our own recent laser microdissection-assisted RNA profiling of motor neurons isolated from two different SOD1 mutant ALS mouse lines (SOD1 G37R and SOD1…”

Section: Significancementioning

confidence: 99%

C1q induction and global complement pathway activation do not contribute to ALS toxicity in mutant SOD1 mice

Lobsiger

Boillée

Pozniak³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Accumulating evidence from mice expressing ALS-causing mutations in superoxide dismutase (SOD1) has implicated pathological immune responses in motor neuron degeneration. This includes microglial activation, lymphocyte infiltration, and the induction of C1q, the initiating component of the classic complement system that is the protein-based arm of the innate immune response, in motor neurons of multiple ALS mouse models expressing dismutase active or inactive SOD1 mutants. Robust induction early in disease course is now identified for multiple complement components (including C1q, C4, and C3) in spinal cords of SOD1 mutantexpressing mice, consistent with initial intraneuronal C1q induction, followed by global activation of the complement pathway. We now test if this activation is a mechanistic contributor to disease. Deletion of the C1q gene in mice expressing an ALS-causing mutant in SOD1 to eliminate C1q induction, and complement cascade activation that follows from it, is demonstrated to produce changes in microglial morphology accompanied by enhanced loss, not retention, of synaptic densities during disease. C1q-dependent synaptic loss is shown to be especially prominent for cholinergic C-bouton nerve terminal input onto motor neurons in affected C1q-deleted SOD1 mutant mice. Nevertheless, overall onset and progression of disease are unaffected in C1q-and C3-deleted ALS mice, thus establishing that C1q induction and classic or alternative complement pathway activation do not contribute significantly to SOD1 mutant-mediated ALS pathogenesis in mice.amyotrophic lateral sclerosis | motoneuron | synaptic density | gender differences | neuroinflammation

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Sporadic ALS has compartment-specific aberrant exon splicing and altered cell–matrix adhesion biology

Cited by 123 publications

References 76 publications

ALS-associated mutation FUS-R521C causes DNA damage and RNA splicing defects

ALS-associated mutation FUS-R521C causes DNA damage and RNA splicing defects

fus/TLS orchestrates splicing of developmental regulators during gastrulation

C1q induction and global complement pathway activation do not contribute to ALS toxicity in mutant SOD1 mice

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