Improved detection of RNA foci in C9orf72 amyotrophic lateral sclerosis post-mortem tissue using BaseScope™ shows a lack of association with cognitive dysfunction

Acknowledgements The authors would like to thank: (i) the Medical Research Council Edinburgh Brain Bank (ethics approval from East of Scotland Research Ethics Service, 16/ES/0084)) for supplying all post-mortem brain material and the Scottish Motor Neurone Disease Register (SMNDR)/Care Audit Research and Evaluation for Motor Neurone Disease (CARE-MND) consortium for all clinical and demographic data (ethical approval from Scotland A Research Ethics Committee 10/MRE00/78 and 15/SS/0216); (ii) The Scottish MND Clinical Specialist team for obtaining consent from patients with MND for inclusion in these studies; (iii) MND Scotland and the Sylvia Aitken Charitable Trust for funding CS to help to establish the MND Tissue Bank. The authors would also like to thank Advanced Cell Diagnostics for gifting the C9orf72 probe (BaseScopeTM Probe - BA-GGGGCCn-3zz-st, Cat Code: 704181), prior to it being made commercially available. Funding A.R.M. is a Lady Edith Wolfson Clinical Fellow and is jointly funded by the Medical Research Council and the Motor Neurone Disease Association (MR/R001162/1). He also acknowledges support from the Rowling Scholars scheme, administered by the Anne Rowling Regenerative Neurology Clinic, University of Edinburgh, Edinburgh, UK. The Chandran laboratory is supported by the Euan MacDonald Centre and the UK Dementia Research Institute, which receives its funding from UK DRI Ltd, funded by the UK Medical Research Council, Alzheimer’s Society and Alzheimer’s Research UK. J.M.G. is funded by a starter grant for clinical lecturers from the Academy of Medical Sciences (AMS: 210JMG 3102 R45620) and C.S. is supported by a Medical Research Council grant (MR/L016400/1).Peer reviewe

Senior Recital: Josh Inglis, saxophone

This recital is presented in partial fulfillment of requirements for the degrees Bachelor of Music in Performance and Bachelor of Music in Music Education. Mr. Inglis studies saxophone with Sam Skelton.https://digitalcommons.kennesaw.edu/musicprograms/2221/thumbnail.jp

FUS-SMN Protein Interactions Link the Motor Neuron Diseases ALS and SMA

SummaryMutations in the RNA binding protein FUS cause amyotrophic lateral sclerosis (ALS), a fatal adult motor neuron disease. Decreased expression of SMN causes the fatal childhood motor neuron disorder spinal muscular atrophy (SMA). The SMN complex localizes in both the cytoplasm and nuclear Gems, and loss of Gems is a cellular hallmark of fibroblasts in patients with SMA. Here, we report that FUS associates with the SMN complex, mediated by U1 snRNP and by direct interactions between FUS and SMN. Functionally, we show that FUS is required for Gem formation in HeLa cells, and expression of FUS containing a severe ALS-causing mutation (R495X) also results in Gem loss. Strikingly, a reduction in Gems is observed in ALS patient fibroblasts expressing either mutant FUS or TDP-43, another ALS-causing protein that interacts with FUS. The physical and functional interactions among SMN, FUS, TDP-43, and Gems indicate that ALS and SMA share a biochemical pathway, providing strong support for the view that these motor neuron diseases are related

Motor function and behaviour across the ALS-FTD spectrum

BACKGROUND: Behavioural/functional disturbances, characteristic of frontotemporal dementia (FTD), are also a feature of amyotrophic lateral sclerosis (ALS) and patients with combined ALS and FTD (FTD-ALS).  AIM OF THE STUDY: To investigate the progression of behavioural disturbances in ALS and FTD using the frontotemporal dementia functional rating scale (FTDFRS).  METHODS: Patients with ALS, FTD-ALS, and FTD were recruited from specialist clinics. Baseline assessments included the FTDFRS and the amyotrophic lateral sclerosis functional rating scale – revised (ALSFRS-R). Baseline assessments were included, as were longitudinal assessments in a proportion of patients.  RESULTS: In total, 21 ALS, 12 FTD-ALS and 14 behavioural variant FTD (bvFTD) patients were included in the study. Moderate or severe behavioural disturbance was common in ALS patients at baseline (47.6%), although less frequent than in bvFTD patients; FTDALS patients displayed intermediate impairment. The ALSFRS-R showed the opposite pattern and did not correlate with the FTDFRS. During the follow-up period, significant (p<0.05) behaviouraldeterioration was demonstrated in bvFTD and FTD-ALS patients, with a trend for decline in ALS patients (p=0.06).  CONCLUSION: Motor disturbance is the primary marker of disease severity in ALS, but behavioural and functional impairment are common, and may decline independently of motor function. As such, the FTDFRS may provide valuable information in the assessment andmonitoring of ALS

FET proteins TAF15 and EWS are selective markers that distinguish FTLD with FUS pathology from amyotrophic lateral sclerosis with FUS mutations

Accumulation of the DNA/RNA binding protein fused in sarcoma as cytoplasmic inclusions in neurons and glial cells is the pathological hallmark of all patients with amyotrophic lateral sclerosis with mutations in FUS as well as in several subtypes of frontotemporal lobar degeneration, which are not associated with FUS mutations. The mechanisms leading to inclusion formation and fused in sarcoma-associated neurodegeneration are only poorly understood. Because fused in sarcoma belongs to a family of proteins known as FET, which also includes Ewing's sarcoma and TATA-binding protein-associated factor 15, we investigated the potential involvement of these other FET protein family members in the pathogenesis of fused in sarcoma proteinopathies. Immunohistochemical analysis of FET proteins revealed a striking difference among the various conditions, with pathology in amyotrophic lateral sclerosis with FUS mutations being labelled exclusively for fused in sarcoma, whereas fused in sarcoma-positive inclusions in subtypes of frontotemporal lobar degeneration also consistently immunostained for TATA-binding protein-associated factor 15 and variably for Ewing's sarcoma. Immunoblot analysis of proteins extracted from post-mortem tissue of frontotemporal lobar degeneration with fused in sarcoma pathology demonstrated a relative shift of all FET proteins towards insoluble protein fractions, while genetic analysis of the TATA-binding protein-associated factor 15 and Ewing's sarcoma gene did not identify any pathogenic variants. Cell culture experiments replicated the findings of amyotrophic lateral sclerosis with FUS mutations by confirming the absence of TATA-binding protein-associated factor 15 and Ewing's sarcoma alterations upon expression of mutant fused in sarcoma. In contrast, all endogenous FET proteins were recruited into cytoplasmic stress granules upon general inhibition of Transportin-mediated nuclear import, mimicking the findings in frontotemporal lobar degeneration with fused in sarcoma pathology. These results allow a separation of fused in sarcoma proteinopathies caused by FUS mutations from those without a known genetic cause based on neuropathological features. More importantly, our data imply different pathological processes underlying inclusion formation and cell death between both conditions; the pathogenesis in amyotrophic lateral sclerosis with FUS mutations appears to be more restricted to dysfunction of fused in sarcoma, while a more global and complex dysregulation of all FET proteins is involved in the subtypes of frontotemporal lobar degeneration with fused in sarcoma patholog

NetDiff – Bayesian model selection for differential gene regulatory network inference

Differential networks allow us to better understand the changes in cellular processes that are exhibited in conditions of interest, identifying variations in gene regulation or protein interaction between, for example, cases and controls, or in response to external stimuli. Here we present a novel methodology for the inference of differential gene regulatory networks from gene expression microarray data. Specifically we apply a Bayesian model selection approach to compare models of conserved and varying network structure, and use Gaussian graphical models to represent the network structures. We apply a variational inference approach to the learning of Gaussian graphical models of gene regulatory networks, that enables us to perform Bayesian model selection that is significantly more computationally efficient than Markov Chain Monte Carlo approaches. Our method is demonstrated to be more robust than independent analysis of data from multiple conditions when applied to synthetic network data, generating fewer false positive predictions of differential edges. We demonstrate the utility of our approach on real world gene expression microarray data by applying it to existing data from amyotrophic lateral sclerosis cases with and without mutations in C9orf72, and controls, where we are able to identify differential network interactions for further investigation

The neural correlates and clinical characteristics of psychosis in the frontotemporal dementia continuum and the C9orf72 expansion

Objective: This present study aims to address the gap in the literature regarding the severity and underlying neural correlates of psychotic symptoms in frontotemporal dementia with and without the C9orf72 gene expansion. Methods: Fifty-six patients with behavioural variant frontotemporal dementia (20 with concomitant amyotrophic lateral sclerosis) and 23 healthy controls underwent neuropsychological assessments, detailed clinical interview for assessment of psychosis symptoms, brain MRI and genetic testing. Carers underwent a clinical interview based upon the neuropsychiatric inventory. Patients were assessed at ForeFront, the Frontotemporal Dementia Research Group at Neuroscience Research Australia or at the Brain and Mind Centre, between January 2008 and December 2013. An index of psychosis was calculated, taking into account the degree and severity of psychosis in each case. Voxel-based morphometry analyses were used to explore relationships between the psychosis index and grey matter changes. Results: Thirty-four percent of frontotemporal dementia patients showed psychotic features. C9orf72 expansion cases were more likely to exhibit psychotic symptoms than non-carriers (64% vs. 26%; p = 0.006), which were also more severe (psychotic index 23.1 vs. 8.1; p = 0.002). Delusions comprised persecutory, somatic, jealous and grandiose types and were present in 57% of C9orf72 carriers and 19% of non-carriers (p = 0.006). Auditory, visual or tactile hallucinations were present in 36% of C9orf72 carriers and 17% of non-carriers (p = 0.13). Increased psychotic symptoms in C9orf72 expansion carriers correlated with atrophy in a distributed cortical and subcortical network that included discrete regions of the frontal, temporal and occipital cortices, as well as the thalamus, striatum and cerebellum. Conclusions: This study underlines the need to consider and assess for psychotic symptoms in the frontotemporal dementia-amyotrophic lateral sclerosis continuum particularly in those with C9orf72 gene expansions. The network of brain regions identified in this study is strikingly similar to that identified in other psychotic disorders such as schizophrenia, which suggests that treatment strategies in psychiatry may be beneficial for the management of psychotic symptoms in frontotemporal dementia

Improved PCR based methods for detecting C9orf72 hexanucleotide repeat expansions

Due to the GC-rich, repetitive nature of C9orf72 hexanucleotide repeat expansions, PCR based detection methods are challenging. Several limitations of PCR have been reported and overcoming these could help to define the pathogenic range. There is also a need to develop improved repeat-primed PCR assays which allow detection even in the presence of genomic variation around the repeat region. We have optimised PCR conditions for the C9orf72 hexanucleotide repeat expansion, using betaine as a co-solvent and specific cycling conditions, including slow ramping and a high denaturation temperature. We have developed a flanking assay, and repeat-primed PCR assays for both 3′ and 5′ ends of the repeat expansion, which when used together provide a robust strategy for detecting the presence or absence of expansions greater than ∼100 repeats, even in the presence of genomic variability at the 3′ end of the repeat. Using our assays, we have detected repeat expansions in 47/442 Scottish ALS patients. Furthermore, we recommend the combined use of these assays in a clinical diagnostic setting

Neuroimaging signatures of frontotemporal dementia genetics: C9ORF72, tau, progranulin and sporadics

A major recent discovery was the identification of an expansion of a non-coding GGGGCC hexanucleotide repeat in the C9ORF72 gene in patients with frontotemporal dementia and amyotrophic lateral sclerosis. Mutations in two other genes are known to account for familial frontotemporal dementia: microtubule-associated protein tau and progranulin. Although imaging features have been previously reported in subjects with mutations in tau and progranulin, no imaging features have been published in C9ORF72. Furthermore, it remains unknown whether there are differences in atrophy patterns across these mutations, and whether regional differences could help differentiate C9ORF72 from the other two mutations at the single-subject level. We aimed to determine the regional pattern of brain atrophy associated with the C9ORF72 gene mutation, and to determine which regions best differentiate C9ORF72 from subjects with mutations in tau and progranulin, and from sporadic frontotemporal dementia. A total of 76 subjects, including 56 with a clinical diagnosis of behavioural variant frontotemporal dementia and a mutation in one of these genes (19 with C9ORF72 mutations, 25 with tau mutations and 12 with progranulin mutations) and 20 sporadic subjects with behavioural variant frontotemporal dementia (including 50% with amyotrophic lateral sclerosis), with magnetic resonance imaging were included in this study. Voxel-based morphometry was used to assess and compare patterns of grey matter atrophy. Atlas-based parcellation was performed utilizing the automated anatomical labelling atlas and Statistical Parametric Mapping software to compute volumes of 37 regions of interest. Hemispheric asymmetry was calculated. Penalized multinomial logistic regression was utilized to create a prediction model to discriminate among groups using regional volumes and asymmetry score. Principal component analysis assessed for variance within groups. C9ORF72 was associated with symmetric atrophy predominantly involving dorsolateral, medial and orbitofrontal lobes, with additional loss in anterior temporal lobes, parietal lobes, occipital lobes and cerebellum. In contrast, striking anteromedial temporal atrophy was associated with tau mutations and temporoparietal atrophy was associated with progranulin mutations. The sporadic group was associated with frontal and anterior temporal atrophy. A conservative penalized multinomial logistic regression model identified 14 variables that could accurately classify subjects, including frontal, temporal, parietal, occipital and cerebellum volume. The principal component analysis revealed similar degrees of heterogeneity within all disease groups. Patterns of atrophy therefore differed across subjects with C9ORF72, tau and progranulin mutations and sporadic frontotemporal dementia. Our analysis suggested that imaging has the potential to be useful to help differentiate C9ORF72 from these other groups at the single-subject level