Genetically mediated increased expression of syntaxin-6, a SNARE protein involved in intracellular protein trafficking, is a proposed risk mechanism for progressive supranuclear palsy and sporadic prion disease. Increased syntaxin-6 protein levels are also causally associated with Alzheimer’s disease, suggesting it may have shared roles across multiple neurodegenerative diseases. However, no study has validated its functional role in tauopathies. To validate a role for syntaxin-6 in tauopathy pathogenesis, we knocked out syntaxin-6 in humanised P301S tauopathy mice. Mice underwent longitudinal rotarod testing, gait analysis, frailty and weight assessment, with neuropathological, biochemical and pathological analyses at 3 and 5 months. Stx6+/+;hTauP301S/P301S mice showed motor impairment from 1 month of age, which was partially rescued by syntaxin-6 knockout from months 1 to 4, with additional protection of gait at 5.5 months. Physiologically, syntaxin-6 knockout exerted a protective effect on weight trajectories and measures of frailty. Reduced neurodegeneration in the superficial cortex was observed at 3 months, as well as higher synaptic coverage at 5 months of age, supporting preserved neuropathological measures related to function. We further observed localised increases in tau pathology in the spinal cord and defined brain regions in young Stx6−/−;hTauP301S/P301S mice, despite total tau levels being comparable, in keeping with altered trafficking of pathological tau species with syntaxin-6 knockout. Despite a partial, early phenotypic rescue of functional measures, terminal endpoint comparisons were confounded by a 20

Increased expression of syntaxin-6, a SNARE protein involved in intracellular protein trafficking, is a proposed genetic risk mechanism for sporadic prion disease and progressive supranuclear palsy, as well as being implicated in Alzheimer’s disease. However, no study has validated its functional role in prion disease, its mechanism of action nor explored the disease stage at which it is acting. Here, we show that syntaxin-6 knockdown in cellular models increases cell-associated infectivity, whilst overexpression produces the opposite effect. This observation is broadly consistent across multiple cell types and prion strains. Furthermore, syntaxin-6 knockdown leads to an accumulation of perinuclear disease-related PrP, consistent with a trafficking mechanism, and alters the morphology of disease-related PrP. We demonstrate that syntaxin-6 knockdown reduces the secretion of prions from infected cells, which provides a mechanism for the prion-related cellular phenotypes observed. Complementary in vivo studies showed that syntaxin-6 influences early stages of prion disease in experimental mice, increasing transmission risk after inoculation with low prion doses. Conversely, syntaxin-6 does not affect prion propagation kinetics or toxicity during established disease. Taken together, our studies firmly establish syntaxin-6 as a modifier of prion pathogenesis with a role in prion trafficking and export. Our findings further suggest that syntaxin-6 modifies the risk of the establishment of disease in line with its genetic association in humans. Thus, this work provides important insights into the role of a pleiotropic prion/prion-like modifier, grounded in human genetics evidence, which may have wider relevance to other neurodegenerative diseases.

Syntaxin-6, a SNARE protein involved in intracellular protein trafficking, is a proposed risk factor for sporadic prion disease, progressive supranuclear palsy and Alzheimer's disease. However, no study has validated its functional role in these diseases, explored the disease stage at which it is acting nor its mechanism of action. Here, we show that syntaxin-6 acts at early stages of prion disease in experimental mice by increasing disease transmission risk following inoculation with low prion doses. Conversely, syntaxin-6 does not affect prion propagation kinetics or toxicity during established disease. Syntaxin-6 manipulation in cellular models profoundly alters the subcellular distribution and morphologies of disease-related PrP and modifies prion export. Furthermore, syntaxin-6 knockout in a transgenic tauopathy mouse model exerts protective effects on numerous physiological, behavioural and neuropathological outcome measures. Therefore, our studies firmly establish syntaxin-6 as a modifier of prion and tau pathogenesis, providing key insights into a fundamental mechanism of neurodegeneration. ### Competing Interest Statement J.C. is the director of D-Gen, Ltd., an academic spin-out company working in the field of prion disease diagnosis, decontamination and therapeutics. J.C. and G.S.J. are shareholders of D-Gen. D-Gen supplied the ICSM35 antibody used for PrP immunohistochemistry and ICSM18 antibody used for cell experiments. The other authors declare no competing interests.

AbstractAn autosomal dominant GGGGCC repeat expansion in intron 1 of theC9orf72gene is the most common genetic cause of both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Here, we set out to engineer a gene targeted mouse model harbouring a pathogenic length humanisedC9orf72repeat expansion allele, in order to model pathological mechanisms in a physiological context. In human disease, pathogenic repeats typically range from the hundreds to thousands of units in length, representing a considerable challenge for cellular and in vivo model generation given the instability of GC rich and repetitive DNA sequences during molecular cloning. To overcome this challenge, we developed new methodology to synthetically and iteratively build pure GGGGCC repeats within a linear vector system, which we then seamlessly and scarlessly embedded within the native human genomic sequence. This created a gene targeting DNA vector for homologous recombination of the human sequence in mouse embryonic stem cells. We used this novel targeting vector to generate a new gene targeted mouse allele,C9orf72h370, that for the first time has mouseC9orf72intron 1 scarlessly replaced with human intron 1 including a pure (GGGGCC)370hexanucleotide repeat expansion. We confirm that the mouse model expresses human intron 1-derived RNA and produces dipeptide repeat proteins derived from the GGGGCC repeat expansion. We now provide this model as a new freely available resource for the field. In addition, we demonstrate the utility of our cloning method for engineering diverse repeat expansion sequences for modelling other disorders, such as Fragile X Syndrome.

Amyotrophic lateral sclerosis (ALS) caused by mutation in superoxide dismutase 1 ( SOD1 ) accounts for 15-30% of familial ALS and is typically autosomal dominant. How single base pair/amino acid changes in this small protein cause neurodegeneration is unknown. In North America, SOD1A4V is the most common familial ALS SOD1 mutation and results in an aggressive form of ALS. Here, we present a novel genomically humanised mouse model of SOD1A4V , in which the mouse Sod1 locus has been replaced by the human SOD1 gene, with intact genomic architecture of exons and introns, but bearing an A4V mutation. In agreement with previously reported human genomic knock-in mice, the phenotype is mild; however, transcriptomic and metabolomic profiling reveal significant dysregulation of glycolysis, the tricarboxylic acid (TCA) cycle, and lipid metabolism. These changes suggest an early bioenergetic imbalance that precedes neuromuscular impairment. Our findings support metabolic dysfunction as an early event in ALS pathogenesis. This freely available SOD1A4V model provides a valuable tool for studying ALS progression and identifying therapeutic targets for pre-symptomatic treatment. SUMMARY STATEMENT This study describes the generation and analysis of novel genomically humanised SOD1A4V mice, revealing metabolic dysfunction through integrated multi-omic analyses, characterising a freely available potential pre-symptomatic ALS model for future research. ### Competing Interest Statement The authors have declared no competing interest.

Induced pluripotent stem cell (iPSC) models are powerful tools for neurodegenerative disease modelling, as they allow mechanistic studies in a human genetic environment and they can be differentiated into a range of neuronal and non-neuronal cells. However, these models come with inherent challenges due to line-to-line and clonal variability. To combat this issue, the iPSC Neurodegenerative Disease Initiative (iNDI) has generated an iPSC repository using a single clonal reference line, KOLF2.1J, into which disease-causing mutations and revertants are introduced via gene editing. Here we describe the generation and validation of lines carrying the most common causative mutation for amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), a repeat expansion in the C9orf72 gene, for the iNDI collection of neurodegenerative iPSC models. We demonstrate that these C9orf72 knock-in lines differentiate efficiently into neurons and display characteristic C9orf72-associated pathologies, including reduced C9orf72 levels and the presence of dipeptide repeat proteins (DPRs) and RNA foci, which increase in abundance over time in culture. These pathologies are not present in revertant cells lacking the repeat expansion. These repeat expansion and revertant cell lines are now available to academic and for-profit institutions through the JAX iPS cell repository and will help to facilitate and standardise iPSC-based ALS/FTD research.

TDP-43 is a nuclear protein encoded by the TARDBP gene, which forms pathological aggregates in various neurodegenerative diseases, collectively known as TDP-43 proteinopathies, including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). These diseases are characterized by multiple pathological mechanisms, with disruptions in lipid regulatory pathways emerging as a critical factor. However, the role of TDP-43 in the regulation of the brain lipid homeostasis and the potential connection of TDP-43 dysfunction to myelin alterations in TDP-43 proteionopathies remain poorly understood, despite the fact that lipids, particularly cholesterol, comprise nearly 70% of myelin. To investigate the causal relationship between TDP-43 dysfunction and disruptions in brain cholesterol homeostasis, we conducted multi-omics analyses (lipidomics, transcriptomics, and functional splicing) on the frontal cortex from the TardbpM323K/M323K knock-in mouse model. Lipidomic analysis revealed alterations in lipid pathways related to membrane composition and lipid droplet accumulation, particularly affecting cholesterol-related species. We found higher lipid droplet accumulation in primary fibroblasts derived from these mice, as well as in the brain of the mutant mice. Similarly, the immunohistochemical detection of a lipid droplet marker was higher in the postmortem frontal cortex, gray matter, and white matter of FTLD-TDP patients compared to non-neurological controls. Transcriptomic analyses showed that TDP-43 pathology led to transcriptional dysregulation of genes essential for myelin production and maintenance. We identified impaired cholesterol metabolism, mainly through the downregulation of endogenous cholesterol synthesis, alongside upregulated cholesterol transport pathways, which we further replicated in FTLD-TDP patients transcriptomic datasets. Collectively, our findings suggest that TDP-43 dysfunction disrupts brain cholesterol homeostasis, potentially compromising myelin integrity.

Background The pathogenic G4C2 repeat expansion in the C9ORF72 gene is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Studies focused on delineating the underlying perturbed mechanisms resulting from this genetic mutation are often confounded by the heterogeneity present in current disease models, such as patient-derived iPSC lines, with estimations of up to 50% of the variation in iPSC cell phenotypes resulting from inter-individual differences. Isogenic models, in which the pathogenic mutation is introduced into a defined genetic background, offer a powerful approach to isolating mutation-specific effects and enable high-resolution comparison across distinct ALS/FTD-associated mutations. Such models are essential for uncovering convergent disease mechanisms and improving reproducibility in ALS/FTD research. Methods A two-step scarless CRISPR/Cas9 genome editing strategy was used to generate isogenic human iPSC lines carrying a de novo knock-in of a disease-length G4C2 repeat expansion in the C9ORF72 locus. The resulting lines underwent thorough quality control and were differentiated into lower motor neurons and assessed for the presence of key ALS/FTD pathologies, including changes to C9ORF72 mRNA and protein expression, RNA foci and dipeptide repeat proteins. Results Two C9ORF72 knock-in iPSC lines were generated with 631 and 600 G4C2 repeats, alongside an isogenic genome editing control line. The C9ORF72 G4C2 repeat expansion knock-in iPSC lines exhibit both loss-of-function and gain-of-function pathological features characteristic of ALS/FTD. Compared to the parental wild-type KOLF2.1J line and isogenic (wild-type) CRISPR control line, these exhibit a significant reduction in C9ORF72 mRNA and protein levels, the presence of RNA foci accumulation, and a marked increase in poly(GA) and poly(GP) dipeptide repeat protein levels in iPSCs and motor neurons. Conclusions This is one of the first reports of a successful knock-in of the pathogenic C9ORF72 G4C2 repeat expansion into a human iPSC line, establishing a genetically defined and physiologically relevant model of ALS/FTD. These isogenic lines recapitulate both key loss- and gain-of-function disease pathologies, providing a crucial complement to existing patient-derived iPSC banks. By eliminating confounding genetic background variability, these cell lines will enable more precise interrogation of C9ORF72 -linked pathomechanisms and offer a robust platform for comparative studies across the ALS and FTD spectrum, mechanistic investigations, and future therapeutic targeting with enhanced translational relevance. ### Competing Interest Statement The authors have declared no competing interest. * ALS : amyotrophic lateral sclerosis ASO : antisense oligonucleotide DPR : dipeptide repeat FTD : frontotemporal dementia HRE : hexanucleotide repeat iPSC : induced pluripotent stem cell liMNs : lower induced motor neurons RAN : repeat associated non-ATG translation rpPCR : repeat primed PCR ssODN : single-stranded oligodeoxyribonucleotide Motor Neurone Disease Association, https://ror.org/02gq0fg61, Ruepp/Apr19/872-791 UK Dementia Research Institute, https://ror.org/02wedp412, UK DRI-6204, UK DRI-6203, UK DRI 1203

The epizootic prion disease of cattle, bovine spongiform encephalopathy (BSE), caused variant Creutzfeldt-Jakob disease (vCJD) in humans following dietary exposure. Codon 129 polymorphism of the human prion protein gene (PRNP), encoding either methionine (M) or valine (V), dictates the propagation of distinct human prion strains and up to now all but one neuropathologically confirmed vCJD patients have had a 129MM genotype. Concordant with this genetic association, transgenic modelling has established that human PrP 129V is incompatible with the vCJD prion strain and that depending on codon 129 genotype, primary human infection with BSE prions may, in addition to vCJD, result in sporadic CJD-like or novel phenotypes. In 2016 we saw the first neuropathologically confirmed case of vCJD in a patient with a codon 129MV genotype. This patient's neuropathology and molecular strain type were pathognomonic of vCJD but their clinical presentation and neuroradiological features were more typical of sporadic CJD, suggestive of possible co-propagation of another prion strain. Here we report the transmission properties of prions from the brain and lymphoreticular tissues of the 129MV vCJD patient. Primary transmissions into transgenic mice expressing human PrP with different codon 129 genotypes mainly produced neuropathological and molecular phenotypes congruent to those observed in the same lines of mice challenged with prions from 129MM vCJD patient brain, indicative that the vCJD prion strain was the dominant propagating prion strain in the patient's brain. Remarkably however, some transgenic mice challenged with 129MV vCJD patient brain propagated a novel prion strain type which at secondary passage was uniformly lethal in mice of all three PRNP codon 129 genotypes after similar short mean incubation periods. These findings establish that cattle BSE prions can trigger the co-propagation of distinct prion strains in humans.

Significant genetic, behavioural and neuroanatomic heterogeneity is common in autism spectrum- and related- neurodevelopmental disorders (NDDs). This heterogeneity constrains the development of effective therapies for diverse patients in precision medicine paradigms. This has led to the search for subgroups of individuals having common etiologic factors/biology (e.g., genetic pathways), thus creating potential uniformity in prognosis and/or treatment response. Despite NDDs having a strong genetic component, only ~15-20% of individuals will present with a specific rare genetic variant considered clinically pathogenic, and therefore, subtyping efforts tend to focus on using clinical, cognitive, and/or brain imaging phenotypes to group individuals. Here we delineated mechanisms via mouse to human translational neuroscience. Using MRI derived structural neuroanatomy and a spatial transcriptomic comparison, we linked subgroups of 135 NDD relevant mouse models (3,515 individual mice) separately to two human databases, with 1,234 and 1,015 human individuals with NDDs, composed of autism, attention-deficit/hyperactivity disorder (ADHD), obsessive compulsive disorder (OCD), other related NDDs, and typically developing controls. Subgroups were significantly linked by consistent neuroanatomy across all three datasets, mouse and human, indicating that direct cross-species subgrouping and translation is consistent and reproducible. Ultimately, four specific neuroanatomical clusters were found and linked to precise molecular mechanisms: two showing a chromatin/transcription motif, with one of those showing specific links to G-protein coupled receptors (GPCR) and Notch signalling, and another two being mainly synaptic in origin, with one off those showing specific connections to axon guidance and Wnt signaling. Assigning molecular pathways, and thus genetic information, from the mouse to individual participants provides an insight into undetected and/or related genetic variants that could be working in combination or interacting with an environmental influence. Moreover, the subgroups found are transdiagnostic, including participants with autism, ADHD, and OCD, which indicates that NDDs as a whole can be subdivided into consistent neuroanatomical clusters with cohesive underlying biological mechanisms. This work allows us to bridge the gap between preclinical models and human disorders, linking previously idiopathic human patients to pertinent genetics, molecular mechanisms, and pathways.

Mutations in the RNA-binding protein FUS lead to nuclear depletion and cytoplasmic mislocalisation of the protein and cause amyotrophic lateral sclerosis (ALS). Using a novel FUS-ALS mouse model, we found that adult mutant mice develop loss of function transcriptomic alterations, along with aberrant cytoplasmic partitioning associated with translatome deficits. Neuromuscular junction innervation was selectively impaired in FUS-ALS females; however, reinnervation following sciatic nerve crush was equally perturbed in both sexes. Additionally, we observed cargo-specific axonal transport alterations, a process critical for neuronal maintenance. In vivo mitochondrial transport was impaired across FUS-ALS mice, whereas the transport of signalling endosomes was selectively disrupted in mutant females. Altogether, our findings identify broader and sex-dependent motor neuron dysfunction in FUS-ALS, emphasise the link between endosomal transport impairments and denervation in disease, and establish FUS-ALS mice as a valuable model for investigating early cellular impairments driving ALS pathology. ### Competing Interest Statement The authors have declared no competing interest. Medical Research Council, https://ror.org/03x94j517, MR/M008606/1, MR/S006508/1, MC\_EX\_MR/N501931/1, MR/Y010949/1, MR/X502984/1 Motor Neurone Disease Association, https://ror.org/02gq0fg61, Birsa/Oct21/976-799, Tosolini/Oct20/973-799 Motor Neuron Disease Research Australia, IG 2450 FightMND Drug Development Grant, DDG-73 Wellcome Trust, https://ror.org/029chgv08, 107116/Z/15/Z, 223022/Z/21/Z UK Dementia Research Institute, UKDRI-1005 China Scholarship Council, https://ror.org/04atp4p48

An autosomal dominant GGGGCC repeat expansion in intron 1 of the C9orf72 gene is the most common genetic cause of both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Here, we set out to engineer a gene targeted mouse model harbouring a pathogenic length humanised C9orf72 repeat expansion allele, in order to model pathological mechanisms in a physiological context. In human disease, pathogenic repeats typically range from the hundreds to thousands of units in length, representing a considerable challenge for cellular and in vivo model generation given the instability of GC rich and repetitive DNA sequences during molecular cloning. To overcome this challenge, we developed new methodology to synthetically and iteratively build pure GGGGCC repeats within a linear vector system, which we then seamlessly and scarlessly embedded within the native human genomic sequence. This created a gene targeting DNA vector for homologous recombination of the human sequence in mouse embryonic stem cells. We used this novel targeting vector to generate a new gene targeted mouse allele, C9orf72h370 , that for the first time has mouse C9orf72 intron 1 scarlessly replaced with human intron 1 including a pure (GGGGCC)370 hexanucleotide repeat expansion. We confirm that the mouse model expresses human intron 1-derived RNA and produces dipeptide repeat proteins derived from the GGGGCC repeat expansion. We now provide this model as a new freely available resource for the field. In addition, we demonstrate the utility of our cloning method for engineering diverse repeat expansion sequences for modelling other disorders, such as Fragile X Syndrome. ### Competing Interest Statement The authors have declared no competing interest.

TDP-43 pathology is found in several neurodegenerative disorders, collectively referred to as “TDP-43 proteinopathies”. Aggregates of TDP-43 are present in the brains and spinal cords of >97% of amyotrophic lateral sclerosis (ALS), and in brains of ∼50% of frontotemporal dementia (FTD) patients. While mutations in the TDP-43 gene ( TARDBP ) are usually associated with ALS, many clinical reports have linked these mutations to cognitive impairments and/or FTD, but also to other neurodegenerative disorders including Parkinsonism (PD) or progressive supranuclear palsy (PSP). TDP-43 is a ubiquitously expressed, highly conserved RNA-binding protein that is involved in many cellular processes, mainly RNA metabolism. To investigate systemic pathological mechanisms in TDP-43 proteinopathies, aiming to capture the pleiotropic effects of TDP-43 mutations, we have further characterised a mouse model carrying a point mutation (M323K) within the endogenous Tardbp gene. Homozygous mutant mice developed cognitive and behavioural deficits as early as 3 months of age. This was coupled with significant brain structural abnormalities, mainly in the cortex, hippocampus, and white matter fibres, together with progressive cortical interneuron degeneration and neuroinflammation. At the motor level, progressive phenotypes appeared around 6 months of age. Thus, cognitive phenotypes appeared to be of a developmental origin with a mild associated progressive neurodegeneration, while the motor and neuromuscular phenotypes seemed neurodegenerative, underlined by a progressive loss of upper and lower motor neurons as well as distal denervation. This is accompanied by progressive elevated TDP-43 protein and mRNA levels in cortex and spinal cord of homozygous mutant mice from 3 months of age, together with increased cytoplasmic TDP-43 mislocalisation in cortex, hippocampus, hypothalamus, and spinal cord at 12 months of age. In conclusion, we find that Tardbp M323K homozygous mutant mice model many aspects of human TDP-43 proteinopathies, evidencing a dual role for TDP-43 in brain morphogenesis as well as in the maintenance of the motor system, making them an ideal in vivo model system to study the complex biology of TDP-43.

The TATA box-binding protein-associated factor 1 (TAF1) is a ubiquitously expressed protein and the largest subunit of the basal transcription factor TFIID, which plays a key role in initiation of RNA polymerase II-dependent transcription. TAF1 missense variants in human males cause X-linked intellectual disability, a neurodevelopmental disorder, and TAF1 is dysregulated in X-linked dystonia-parkinsonism, a neurodegenerative disorder. However, this field has lacked a genetic mouse model of TAF1 disease to explore its mechanism in mammals and treatments. Here, we generated and validated a conditional cre-lox allele and the first ubiquitous Taf1 knockout mouse. We discovered that Taf1 deletion in male mice was embryonically lethal, which may explain why no null variants have been identified in humans. In the brains of Taf1 heterozygous female mice, no differences were found in gross structure, overall expression and protein localisation, suggesting extreme skewed X inactivation towards the non-mutant chromosome. Nevertheless, these female mice exhibited a significant increase in weight, weight with age, and reduced movement, suggesting that a small subset of neurons was negatively impacted by Taf1 loss. Finally, this new mouse model may be a future platform for the development of TAF1 disease therapeutics.

Loss of function of the RNA-binding protein TDP-43 (TDP-LOF) is a hallmark of amyotrophic lateral sclerosis (ALS) and other neurodegenerative disorders. Here we describe TDP-REG, which exploits the specificity of cryptic splicing induced by TDP-LOF to drive protein expression when and where the disease process occurs. The SpliceNouveau algorithm combines deep learning with rational design to generate customizable cryptic splicing events within protein-coding sequences. We demonstrate that expression of TDP-REG reporters is tightly coupled to TDP-LOF in vitro and in vivo. TDP-REG enables genomic prime editing to ablate the UNC13A cryptic donor splice site specifically upon TDP-LOF. Finally, we design TDP-REG vectors encoding a TDP-43/Raver1 fusion protein that rescues key pathological cryptic splicing events, paving the way for the development of precision therapies for TDP43-related disorders.

Sporadic Creutzfeldt-Jakob disease (sCJD), the most common human prion disease, is thought to occur when the cellular prion protein (PrPC) spontaneously misfolds and assembles into prion fibrils, culminating in fatal neurodegeneration. In a genome-wide association study of sCJD, we recently identified risk variants in and around the gene STX6, with evidence to suggest a causal increase of STX6 expression in disease-relevant brain regions. STX6 encodes syntaxin-6, a SNARE protein primarily involved in early endosome to trans-Golgi network retrograde transport. Here we developed and characterised a mouse model with genetic depletion of Stx6 and investigated a causal role of Stx6 expression in mouse prion disease through a classical prion transmission study, assessing the impact of homozygous and heterozygous syntaxin-6 knockout on disease incubation periods and prion-related neuropathology. Following inoculation with RML prions, incubation periods in Stx6−/− and Stx6+/− mice differed by 12 days relative to wildtype. Similarly, in Stx6−/− mice, disease incubation periods following inoculation with ME7 prions also differed by 12 days. Histopathological analysis revealed a modest increase in astrogliosis in ME7-inoculated Stx6−/− animals and a variable effect of Stx6 expression on microglia activation, however no differences in neuronal loss, spongiform change or PrP deposition were observed at endpoint. Importantly, Stx6−/− mice are viable and fertile with no gross impairments on a range of neurological, biochemical, histological and skeletal structure tests. Our results provide some support for a pathological role of Stx6 expression in prion disease, which warrants further investigation in the context of prion disease but also other neurodegenerative diseases considering syntaxin-6 appears to have pleiotropic risk effects in progressive supranuclear palsy and Alzheimer's disease.

ABSTRACTFUS (Fused in sarcoma) is a ubiquitously expressed DNA/RNA binding protein. Mutations in FUS cause aggressive juvenile forms of amyotrophic lateral sclerosis (ALS), as in the case with the FUSDelta14 mutation. While most studies have focused on the role of FUS in motor neuron degeneration, little is known about the effect ofFUSmutations in the whole body, and the impact ofFUSmutations in the correct development of the nervous system. We studied pleiotropic phenotypes in a physiological knock-in mouse model carrying the FUSDelta14 mutation in homozygosity. RNA sequencing was conducting in six different tissues (frontal cortex, spinal cord, tibialis anterior muscle, white and brown adipose tissue and liver) to identify the genes and pathways altered by the FUSDelta14 mutant protein in the systemic transcriptome. Additionally, brain structural magnetic resonance imaging (MRI) and histological characterisation was conducted in young mice to study the role of FUS mutation in the brain development. FUS mutant protein was upregulated and mislocalised in the cytoplasm in most cells of the tissues analysed. We identified few genes commonly altered in all tissues by this mutation, although most genes and pathways affected were generally tissue-specific. Phenotypic assessment of mice revealed systemic metabolic alterations related to the pathway changes identified. MRI brain scans revealed that homozygous FUSDelta14 brains were smaller and displayed significant morphological alterations including a thinner cortex, reduced neuronal number and increased gliosis, which correlated with early cognitive impairment and fatal seizures. We demonstrated that the disease aetiology of FUS mutations can include neurodevelopmental and systemic alterations, which should be taken into consideration in the clinic.

Amyotrophic lateral sclerosis is a complex disorder most of which is 'sporadic' of unknown origin but approximately 10% is familial, arising from single mutations in any of more than 30 genes. Thus, there are more than 30 familial ALS subtypes, with different, often unknown, molecular pathologies leading to a complex constellation of clinical phenotypes. We have mouse models for many genetic forms of the disorder, but these do not, on their own, necessarily show us the key pathological pathways at work in human patients. To date, we have no models for the 90% of ALS that is 'sporadic'. Potential therapies have been developed mainly using a limited set of mouse models, and through lack of alternatives, in the past these have been tested on patients regardless of aetiology. Cancer researchers have undertaken therapy development with similar challenges; they have responded by producing complex mouse models that have transformed understanding of pathological processes, and they have implemented patient stratification in multi-centre trials, leading to the effective translation of basic research findings to the clinic. ALS researchers have successfully adopted this combined approach, and now to increase our understanding of key disease pathologies, and our rate of progress for moving from mouse models to mechanism to ALS therapies we need more, innovative, complex mouse models to address specific questions.