Structural basis for mutation-induced destabilization of profilin 1 in ALS

Mutations in profilin 1 (PFN1) are associated with amyotrophic lateral sclerosis (ALS); however, the pathological mechanism of PFN1 in this fatal disease is unknown. We demonstrate that ALS-linked mutations severely destabilize the native conformation of PFN1 in vitro and cause accelerated turnover of the PFN1 protein in cells. This mutation-induced destabilization can account for the high propensity of ALS-linked variants to aggregate and also provides rationale for their reported loss-of-function phenotypes in cell-based assays. The source of this destabilization is illuminated by the X-ray crystal structures of several PFN1 proteins, revealing an expanded cavity near the protein core of the destabilized M114T variant. In contrast, the E117G mutation only modestly perturbs the structure and stability of PFN1, an observation that reconciles the occurrence of this mutation in the control population. These findings suggest that a destabilized form of PFN1 underlies PFN1-mediated ALS pathogenesis.

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Structural basis for the ARF GAP activity and specificity of the C9orf72 complex

Nature Communications ◽

10.1038/s41467-021-24081-0 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Ming-Yuan Su ◽

Simon A. Fromm ◽

Jonathan Remis ◽

Daniel B. Toso ◽

James H. Hurley

Keyword(s):

Amyotrophic Lateral Sclerosis ◽

Active Site ◽

Binding Pocket ◽

Structural Basis ◽

Specific Function ◽

Loss Of Function ◽

Gtpase Activating Protein ◽

Protein Activity ◽

Mtorc1 Pathway ◽

Lateral Sclerosis

AbstractMutation of C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontal temporal degeneration (FTD), which is attributed to both a gain and loss of function. C9orf72 forms a complex with SMCR8 and WDR41, which was reported to have GTPase activating protein activity toward ARF proteins, RAB8A, and RAB11A. We determined the cryo-EM structure of ARF1-GDP-BeF3- bound to C9orf72:SMCR8:WDR41. The SMCR8longin and C9orf72longin domains form the binding pocket for ARF1. One face of the C9orf72longin domain holds ARF1 in place, while the SMCR8longin positions the catalytic finger Arg147 in the ARF1 active site. Mutations in interfacial residues of ARF1 and C9orf72 reduced or eliminated GAP activity. RAB8A GAP required ~10-fold higher concentrations of the C9orf72 complex than for ARF1. These data support a specific function for the C9orf72 complex as an ARF GAP. The structure also provides a model for the active forms of the longin domain GAPs of FLCN and NPRL2 that regulate the Rag GTPases of the mTORC1 pathway.

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Structural basis for the ARF GAP activity and specificity of the C9orf72 complex

10.1101/2021.04.13.439632 ◽

2021 ◽

Author(s):

Ming-Yuan Su ◽

Simon H Fromm ◽

Jonathan Remis ◽

Daniel Toso ◽

James H Hurley

Keyword(s):

Amyotrophic Lateral Sclerosis ◽

Active Site ◽

Binding Pocket ◽

Structural Basis ◽

Specific Function ◽

Loss Of Function ◽

Gtpase Activating Protein ◽

Protein Activity ◽

Data Support ◽

Lateral Sclerosis

Mutation of C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontal temporal degeneration (FTD), which is attributed to both a gain and loss of function. C9orf72 forms a complex with SMCR8 and WDR41, which was reported to have GTPase activating protein activity toward ARF proteins, RAB8A, and RAB11A. We determined the cryo-EM structure of ARF1-GDP-BeF3- bound to C9orf72:SMCR8:WDR41. The SMCR8longin and C9orf72longin domains form the binding pocket for ARF1. One face of the C9orf72longin domain holds ARF1 in place, while the SMCR8longin positions the catalytic finger Arg147 in the ARF1 active site. Mutations in interfacial residues of ARF1 and C9orf72 reduced or eliminated GAP activity. RAB8A GAP required ~10-fold higher concentrations of the C9orf72 complex than for ARF1. These data support a specific function for the C9orf72 complex as an ARF GAP.

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The Link between VAPB Loss of Function and Amyotrophic Lateral Sclerosis

Cells ◽

10.3390/cells10081865 ◽

2021 ◽

Vol 10 (8) ◽

pp. 1865

Author(s):

Nica Borgese ◽

Nicola Iacomino ◽

Sara Francesca Colombo ◽

Francesca Navone

Keyword(s):

Amyotrophic Lateral Sclerosis ◽

Motor Neurons ◽

Adaptor Proteins ◽

Loss Of Function ◽

Membrane Contact Sites ◽

Physiological Processes ◽

Main Driver ◽

Single Allele ◽

Induced Pluripotent ◽

Lateral Sclerosis

The VAP proteins are integral adaptor proteins of the endoplasmic reticulum (ER) membrane that recruit a myriad of interacting partners to the ER surface. Through these interactions, the VAPs mediate a large number of processes, notably the generation of membrane contact sites between the ER and essentially all other cellular membranes. In 2004, it was discovered that a mutation (p.P56S) in the VAPB paralogue causes a rare form of dominantly inherited familial amyotrophic lateral sclerosis (ALS8). The mutant protein is aggregation-prone, non-functional and unstable, and its expression from a single allele appears to be insufficient to support toxic gain-of-function effects within motor neurons. Instead, loss-of-function of the single wild-type allele is required for pathological effects, and VAPB haploinsufficiency may be the main driver of the disease. In this article, we review the studies on the effects of VAPB deficit in cellular and animal models. Several basic cell physiological processes are affected by downregulation or complete depletion of VAPB, impinging on phosphoinositide homeostasis, Ca2+ signalling, ion transport, neurite extension, and ER stress. In the future, the distinction between the roles of the two VAP paralogues (A and B), as well as studies on motor neurons generated from induced pluripotent stem cells (iPSC) of ALS8 patients will further elucidate the pathogenic basis of p.P56S familial ALS, as well as of other more common forms of the disease.

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Motor neuron disease in vitro: the use of cultured motor neurons to study amyotrophic lateral sclerosis

European Journal of Pharmacology ◽

10.1016/s0014-2999(00)00560-4 ◽

2000 ◽

Vol 405 (1-3) ◽

pp. 285-295 ◽

Cited By ~ 22

Author(s):

P.R Bär

Keyword(s):

Amyotrophic Lateral Sclerosis ◽

Motor Neuron ◽

Motor Neuron Disease ◽

Motor Neurons ◽

Lateral Sclerosis

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“My World Has Expanded Even Though I'm Stuck at Home”: Experiences of Individuals With Amyotrophic Lateral Sclerosis Who Use Augmentative and Alternative Communication and Social Media

American Journal of Speech-Language Pathology ◽

10.1044/2015_ajslp-15-0010 ◽

2015 ◽

Vol 24 (4) ◽

pp. 680-695 ◽

Cited By ~ 22

Author(s):

Jessica Caron ◽

Janice Light

Keyword(s):

Social Media ◽

Amyotrophic Lateral Sclerosis ◽

Augmentative And Alternative Communication ◽

Future Research ◽

Loss Of Function ◽

Alternative Communication ◽

Communication Modes ◽

Communication Needs ◽

Use Of Social Media ◽

Lateral Sclerosis

PurposeThis study aimed to expand the current understanding of how persons with amyotrophic lateral sclerosis (pALS) use augmentative and alternative communication and social media to address their communication needs.MethodAn online focus group was used to investigate the experiences of 9 pALS who use augmentative and alternative communication and social media. Questions posed to the group related to (a) current use of social media, (b) advantages of social media, (c) barriers to independent use, (d) supports to independent use, and (e) recommendations for developers, policy makers, and other pALS.ResultsParticipants primarily reported that use of social media was a beneficial tool that provided increased communication opportunities, connections to communication partners, and networks of support. Specific results are discussed with reference to the research as well as implications for practice and recommendations for future research.ConclusionsAs individuals with ALS experience loss of function, some communication modes may no longer be viable. Providing access to different modes of communication, including social media, can allow independence, participation and better quality of life.

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Interarticulator Coordination in Dysarthria

Journal of Speech Language and Hearing Research ◽

10.1044/1092-4388(2003/097) ◽

2003 ◽

Vol 46 (5) ◽

pp. 1247-1261 ◽

Cited By ~ 29

Author(s):

Gary Weismer ◽

Yana Yunusova ◽

John R. Westbury

Keyword(s):

Parkinson’S Disease ◽

Amyotrophic Lateral Sclerosis ◽

Speech Production ◽

Experimental Work ◽

Vowel Duration ◽

Global Synchronization ◽

X Ray ◽

Acoustic Target ◽

Temporal Location ◽

Lateral Sclerosis

Articulatory discoordination is often said to be an important feature of the speech production disorder in dysarthria, but little experimental work has been done to identify and specify the coordination difficulties. The present study evaluated the coordination of labial and lingual gestures for /u/ production in persons with Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), and in control participants. Both tongue backing/raising and reduction of the area enclosed by the lips can produce the characteristic low F2 of /u/. The timing of these articulatory gestures with respect to the acoustic target of a low F2 was inferred from X-ray microbeam data. Pellet motions of the tongue dorsum and lips revealed the timing of the lingual and labial gestures to be strongly linked together (synchronized), predictive of the temporal location of the lowest F2 within the vocalic nucleus, and scaled proportionately to the overall vowel duration in control participants. Somewhat surprisingly, essentially the same findings were obtained in the speakers with dysarthria. These relationships were noisier among the speakers with dysarthria, but the global synchronization patterns applied to all 3 groups. Further analyses revealed the synchronization to be less well defined and more variable across speakers with ALS, as compared to speakers with PD and the controls. Results are discussed relative to concepts of coordination in dysarthria.

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Concomitant gain and loss of function pathomechanisms in C9ORF72 amyotrophic lateral sclerosis

Life Science Alliance ◽

10.26508/lsa.202000764 ◽

2021 ◽

Vol 4 (4) ◽

pp. e202000764

Author(s):

Arun Pal ◽

Benedikt Kretner ◽

Masin Abo-Rady ◽

Hannes Glaβ ◽

Banaja P Dash ◽

...

Keyword(s):

Amyotrophic Lateral Sclerosis ◽

Loss Of Function ◽

Dna Double Strand Breaks ◽

Hexanucleotide Repeat ◽

Rna Foci ◽

Repeat Expansions ◽

Organelle Motility ◽

Trafficking Defects ◽

Induced Pluripotent ◽

Lateral Sclerosis

Intronic hexanucleotide repeat expansions (HREs) in C9ORF72 are the most frequent genetic cause of amyotrophic lateral sclerosis, a devastating, incurable motoneuron (MN) disease. The mechanism by which HREs trigger pathogenesis remains elusive. The discovery of repeat-associated non-ATG (RAN) translation of dipeptide repeat proteins (DPRs) from HREs along with reduced exonic C9ORF72 expression suggests gain of toxic functions (GOFs) through DPRs versus loss of C9ORF72 functions (LOFs). Through multiparametric high-content (HC) live profiling in spinal MNs from induced pluripotent stem cells and comparison to mutant FUS and TDP43, we show that HRE C9ORF72 caused a distinct, later spatiotemporal appearance of mainly proximal axonal organelle motility deficits concomitant to augmented DNA double-strand breaks (DSBs), RNA foci, DPRs, and apoptosis. We show that both GOFs and LOFs were necessary to yield the overall C9ORF72 pathology. Increased RNA foci and DPRs concurred with onset of axon trafficking defects, DSBs, and cell death, although DSB induction itself did not phenocopy C9ORF72 mutants. Interestingly, the majority of LOF-specific DEGs were shared with HRE-mediated GOF DEGs. Finally, C9ORF72 LOF was sufficient—albeit to a smaller extent—to induce premature distal axonal trafficking deficits and increased DSBs.

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Ribosome inhibition by C9ORF72-ALS/FTD-associated poly-PR and poly-GR proteins revealed by cryo-EM

10.1101/2020.08.30.274597 ◽

2020 ◽

Author(s):

Anna B. Loveland ◽

Egor Svidritskiy ◽

Denis Susorov ◽

Soojin Lee ◽

Alexander Park ◽

...

Keyword(s):

Amyotrophic Lateral Sclerosis ◽

Specific Binding ◽

Structural Basis ◽

Transferase Activity ◽

Peptidyl Transferase ◽

Peptidyl Transferase Center ◽

Ribosomal Tunnel ◽

Cellular Translation ◽

Lateral Sclerosis ◽

Dipeptide Repeat

AbstractToxic dipeptide repeat (DPR) proteins are produced from expanded G4C2 hexanucleotide repeats in the C9ORF72 gene, which cause amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Two DPR proteins, poly-PR and poly-GR, repress cellular translation but the molecular mechanism remains unknown. Here we show that poly-PR and poly-GR of ≥ 20 repeats inhibit the ribosome’s peptidyl-transferase activity at nanomolar concentrations, comparable to specific translation inhibitors. High-resolution cryo-EM structures reveal that poly-PR and poly-GR block the polypeptide tunnel of the ribosome, extending into the peptidyl-transferase center. Consistent with these findings, the macrolide erythromycin, which binds in the tunnel, competes with the DPR proteins and restores peptidyl-transferase activity. Our results demonstrate that strong and specific binding of poly-PR and poly-GR in the ribosomal tunnel blocks translation, revealing the structural basis of their toxicity in C9ORF72-ALS/FTD.

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Bisperoxovanadium promotes motor neuron survival and neuromuscular innervation in amyotrophic lateral sclerosis

Molecular Brain ◽

10.1186/s13041-021-00867-7 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Junmei Wang ◽

Lydia Tierney ◽

Ranjeet Mann ◽

Thomas Lonsway ◽

Chandler L. Walker

Keyword(s):

Amyotrophic Lateral Sclerosis ◽

Motor Neuron ◽

Lumbar Spinal Cord ◽

Therapeutic Effects ◽

Spinal Cord Tissue ◽

Neuron Loss ◽

Neuroprotective Therapy ◽

Motor Neuron Loss ◽

Lateral Sclerosis

AbstractAmyotrophic lateral sclerosis (ALS) is the most common motor neuron (MN) disease, with no present cure. The progressive loss of MNs is the hallmark of ALS. We have previously shown the therapeutic effects of the phosphatase and tensin homolog (PTEN) inhibitor, potassium bisperoxo (picolinato) vanadium (bpV[pic]), in models of neurological injury and demonstrated significant neuroprotective effects on MN survival. However, accumulating evidence suggests PTEN is detrimental for MN survival in ALS. Therefore, we hypothesized that treating the mutant superoxide dismutase 1 G93A (mSOD1G93A) mouse model of ALS during motor neuron degeneration and an in vitro model of mSOD1G93A motor neuron injury with bpV(pic) would prevent motor neuron loss. To test our hypothesis, we treated mSOD1G93A mice intraperitoneally daily with 400 μg/kg bpV(pic) from 70 to 90 days of age. Immunolabeled MNs and microglial reactivity were analyzed in lumbar spinal cord tissue, and bpV(pic) treatment significantly ameliorated ventral horn motor neuron loss in mSOD1G93A mice (p = 0.003) while not significantly altering microglial reactivity (p = 0.701). Treatment with bpV(pic) also significantly increased neuromuscular innervation (p = 0.018) but did not affect muscle atrophy. We also cultured motor neuron-like NSC-34 cells transfected with a plasmid to overexpress mutant SOD1G93A and starved them in serum-free medium for 24 h with and without bpV(pic) and downstream inhibitor of Akt signaling, LY294002. In vitro, bpV(pic) improved neuronal viability, and Akt inhibition reversed this protective effect (p < 0.05). In conclusion, our study indicates systemic bpV(pic) treatment could be a valuable neuroprotective therapy for ALS.

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