Nucleolar stress in C9orf72 and sporadic ALS spinal motor neurons precedes TDP-43 mislocalization

AbstractNucleolar stress has been implicated in the pathology and disease pathogenesis of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) from repeat expansions of GGGGCC in C9orf72 (C9-ALS/FTLD) but not in sporadic ALS (SALS). Previously we reported that antisense RNA transcripts are unique in C9-ALS because of their nucleolar localization in spinal motor neurons and correlation with TDP-43 mislocalization, the hallmark proteinopathy of ALS and FTLD. Here we report our further studies of 11 SALS, 11 C9-ALS and 11 control spinal cords. We find that nucleolar stress manifests specifically as shrinkage in nucleoli of C9-ALS spinal motor neurons. Nucleolar size reduction is greatest in similarly sized alpha motor neurons from C9-ALS cases and results are not skewed by the number of surviving neurons from each ALS spinal cord. Surprisingly, nucleolar shrinkage occurs before main pathological hallmarks—TDP-43 mislocalization or antisense RNA foci—appear and this suggest that nucleolar stress can precede pathology in C9-ALS, findings previously identified in C9-FTLD using sense RNA foci and dipeptide repeat proteins as pathological markers. Importantly, these observations are also seen in SALS motor neurons and thus nucleolar stress appears to be a significant and probably upstream problem in sporadic disease.

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Predicting disease specific spinal motor neurons and glia in sporadic ALS

Neurobiology of Disease ◽

10.1016/j.nbd.2019.104523 ◽

2019 ◽

Vol 130 ◽

pp. 104523 ◽

Cited By ~ 3

Author(s):

Fabien Dachet ◽

Jiangou Liu ◽

John Ravits ◽

Fei Song

Keyword(s):

Motor Neurons ◽

Spinal Motor Neurons ◽

Spinal Motor ◽

Sporadic Als ◽

Disease Specific

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Misfolded SOD1 inclusions in patients with mutations in C9orf72 and other ALS/FTD-associated genes

Journal of Neurology Neurosurgery & Psychiatry ◽

10.1136/jnnp-2018-319386 ◽

2019 ◽

Vol 90 (8) ◽

pp. 861-869 ◽

Cited By ~ 30

Author(s):

Karin Forsberg ◽

Karin Graffmo ◽

Bente Pakkenberg ◽

Markus Weber ◽

Martin Nielsen ◽

...

Keyword(s):

Motor Neurons ◽

Superoxide Dismutase 1 ◽

Wild Type ◽

Spinal Motor Neurons ◽

Hexanucleotide Repeat ◽

Cortical Motor ◽

Spinal Motor ◽

Repeat Expansions ◽

Misfolded Sod1 ◽

Lateral Sclerosis

ObjectiveA hallmark of amyotrophic lateral sclerosis (ALS) caused by mutations in superoxide dismutase-1 (SOD1) are inclusions containing SOD1 in motor neurons. Here, we searched for SOD1-positive inclusions in 29 patients carrying ALS-linked mutations in six other genes.MethodsA panel of antibodies that specifically recognise misfolded SOD1 species were used for immunohistochemical investigations of autopsy tissue.ResultsThe 18 patients with hexanucleotide-repeat-expansions in C9orf72 had inclusions of misfolded wild type (WT) SOD1WT in spinal motor neurons. Similar inclusions were occasionally observed in medulla oblongata and in the motor cortex and frontal lobe. Patients with mutations in FUS, KIF5A, NEK1, ALSIN or VAPB, carried similar SOD1WT inclusions. Minute amounts of misSOD1WT inclusions were detected in 2 of 20 patients deceased from non-neurological causes and in 4 of 10 patients with other neurodegenerative diseases. Comparison was made with 17 patients with 9 different SOD1 mutations. Morphologically, the inclusions in patients with mutations in C9orf72HRE, FUS, KIF5A, NEK1, VAPB and ALSIN resembled inclusions in patients carrying the wildtype-like SOD1D90A mutation, whereas patients carrying unstable SOD1 mutations (A4V, V5M, D76Y, D83G, D101G, G114A, G127X, L144F) had larger skein-like SOD1-positive inclusions.Conclusions and relevanceAbundant inclusions containing misfolded SOD1WT are found in spinal and cortical motor neurons in patients carrying mutations in six ALS-causing genes other than SOD1. This suggests that misfolding of SOD1WT can be part of a common downstream event that may be pathogenic. The new anti-SOD1 therapeutics in development may have applications for a broader range of patients.

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Acute neurotoxicant exposure induces hyperexcitability in mouse lumbar spinal motor neurons

Journal of Neurophysiology ◽

10.1152/jn.00775.2019 ◽

2020 ◽

Vol 123 (4) ◽

pp. 1448-1459 ◽

Cited By ~ 2

Author(s):

Michael P. Sceniak ◽

Jake B. Spitsbergen ◽

Shasta L. Sabo ◽

Yukun Yuan ◽

William D. Atchison

Keyword(s):

Spinal Cord ◽

Motor Neurons ◽

Normal Function ◽

Mehg Exposure ◽

Toxicant Exposure ◽

Spinal Motor Neurons ◽

Spinal Motor ◽

Sporadic Als ◽

Lumbar Spinal ◽

Lateral Sclerosis

Spinal motor neurons (MNs) are susceptible to glutamatergic excitotoxicity, an effect associated with lumbar MN degeneration in amyotrophic lateral sclerosis (ALS). MN susceptibility to environmental toxicant exposure, one prospective contributor to sporadic ALS, has not been systematically studied. The goal of this study was to test the ability of a well-known environmental neurotoxicant to induce hyperexcitability in mouse lumbar MNs. Methylmercury (MeHg) causes neurotoxicity through mechanisms involving elevated intracellular Ca2+ concentration ([Ca2+]i), a hallmark of excitotoxicity. We tested whether acute exposure to MeHg induces hyperexcitability in MNs by altering synaptic transmission, using whole cell patch-clamp recordings of lumbar spinal MNs in vitro. Acute MeHg exposure (20 μM) led to an increase in the frequency of both spontaneous excitatory postsynaptic currents (EPSCs) and miniature EPSCs. The frequency of inhibitory postsynaptic currents (IPSCs) was also increased by MeHg. Action potential firing rates, both spontaneous and evoked, were increased by MeHg, despite increases in both EPSCs and IPSCs, indicating a shift toward hyperexcitability. Also consistent with hyperexcitability, fluo 4-AM microfluorimetry indicated that MeHg exposure induced an increase in [Ca2+]i. Spinal cord hyperexcitability is partially mediated by Ca2+-permeable AMPA receptors, as MeHg-dependent increases in EPSCs were blocked by 1-napthyl spermine. Therefore, spinal MNs appear highly susceptible to MeHg exposure, leading to significant increases in spontaneous network excitability and disruption of normal function. Prolonged hyperexcitability could lead to eventual neurodegeneration and loss of motor function as observed in spinal cord after MeHg exposure in vivo and may contribute to MeHg-induced acceleration of ALS symptoms. NEW & NOTEWORTHY Spinal motor neurons (MN) are susceptible to glutamatergic excitotoxicity, an effect associated with lumbar MN degeneration in amyotrophic lateral sclerosis (ALS). This study investigated MN susceptibility to environmental toxicant exposure, one prospective contributor to sporadic ALS. Spinal MNs appear highly susceptible to methylmercury exposure, leading to significant increases in spontaneous network excitability and disruption of normal function. Prolonged hyperexcitability could lead to neurodegeneration and loss of motor function as observed in ALS spinal cord symptoms.

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Novel STMN2 Variant Linked to Amyotrophic Lateral Sclerosis Risk and Clinical Phenotype

Frontiers in Aging Neuroscience ◽

10.3389/fnagi.2021.658226 ◽

2021 ◽

Vol 13 ◽

Author(s):

Frances Theunissen ◽

Ryan S. Anderton ◽

Frank L. Mastaglia ◽

Loren L. Flynn ◽

Samantha J. Winter ◽

...

Keyword(s):

Motor Neurons ◽

Disease Risk ◽

Age Of Onset ◽

Survival Duration ◽

Control Analysis ◽

Ca Repeat ◽

Variable Expression ◽

Spinal Motor Neurons ◽

Spinal Motor ◽

Sporadic Als

ObjectiveThere is a critical need to establish genetic markers that explain the complex phenotypes and pathogenicity of ALS. This study identified a polymorphism in the Stathmin-2 gene and investigated its association with sporadic ALS (sALS) disease risk, age-of onset and survival duration.MethodsThe candidate CA repeat was systematically analyzed using PCR, Sanger sequencing and high throughput capillary separation for genotyping. Stathmin-2 expression was investigated using RT-PCR in patient olfactory neurosphere-derived (ONS) cells and RNA sequencing in laser-captured spinal motor neurons.ResultsIn a case-control analysis of a combined North American sALS cohort (n = 321) and population control group (n = 332), long/long CA genotypes were significantly associated with disease risk (p = 0.042), and most strongly when one allele was a 24 CA repeat (p = 0.0023). In addition, longer CA allele length was associated with earlier age-of-onset (p = 0.039), and shorter survival duration in bulbar-onset cases (p = 0.006). In an Australian longitudinal sALS cohort (n = 67), ALS functional rating scale scores were significantly lower in carriers of the long/long genotype (p = 0.034). Stathmin-2 mRNA expression was reduced in sporadic patient ONS cells. Additionally, sALS patients and controls exhibited variable expression of Stathmin-2 mRNA according to CA genotype in laser-captured spinal motor neurons.ConclusionsWe report a novel non-coding CA repeat in Stathmin-2 which is associated with sALS disease risk and has disease modifying effects. The potential value of this variant as a disease marker and tool for cohort enrichment in clinical trials warrants further investigation.

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