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1.
Int J Mol Sci ; 23(23)2022 Nov 22.
Artículo en Inglés | MEDLINE | ID: mdl-36498882

RESUMEN

Objective markers for the neurodegenerative disorder progressive supranuclear palsy (PSP) are needed to provide a timely diagnosis with greater certainty. Non-coding RNA (ncRNA), including microRNA, piwi-interacting RNA, and transfer RNA, are good candidate markers in other neurodegenerative diseases, but have not been investigated in PSP. Therefore, as proof of principle, we sought to identify whether they were dysregulated in matched serum and cerebrospinal fluid (CSF) samples of patients with PSP. Small RNA-seq was undertaken on serum and CSF samples from healthy controls (n = 20) and patients with PSP (n = 31) in two cohorts, with reverse transcription-quantitative PCR (RT-qPCR) to confirm their dysregulation. Using RT-qPCR, we found in serum significant down-regulation in hsa-miR-92a-3p, hsa-miR-626, hsa-piR-31068, and tRNA-ValCAC. In CSF, both hsa-let-7a-5p and hsa-piR-31068 showed significant up-regulation, consistent with their changes observed in the RNA-seq results. Interestingly, we saw no correlation in the expression of hsa-piR-31068 within our matched serum and CSF samples, suggesting there is no common dysregulatory mechanism between the two biofluids. While these changes were in a small cohort of samples, we have provided novel evidence that ncRNA in biofluids could be possible diagnostic biomarkers for PSP and further work will help to expand this potential.


Asunto(s)
MicroARNs , Parálisis Supranuclear Progresiva , Humanos , Parálisis Supranuclear Progresiva/diagnóstico , Parálisis Supranuclear Progresiva/genética , Biomarcadores , MicroARNs/genética , Regulación hacia Abajo
2.
J Neuroinflammation ; 17(1): 135, 2020 Apr 28.
Artículo en Inglés | MEDLINE | ID: mdl-32345319

RESUMEN

Amyotrophic lateral sclerosis (ALS) is the most common form of motor neuron degeneration in adults, and several mechanisms underlying the disease pathology have been proposed. It has been shown that glia communicate with other cells by releasing extracellular vesicles containing proteins and nucleic acids, including microRNAs (miRNAs), which play a role in the post-transcriptional regulation of gene expression. Dysregulation of miRNAs is commonly observed in ALS patients, together with inflammation and an altered microglial phenotype. However, the role of miRNA-containing vesicles in microglia-to-neuron communication in the context of ALS has not been explored in depth. This review summarises the evidence for the presence of inflammation, pro-inflammatory microglia and dysregulated miRNAs in ALS, then explores how microglia may potentially be responsible for this miRNA dysregulation. The possibility of pro-inflammatory ALS microglia releasing miRNAs which may then enter neuronal cells to contribute to degeneration is also explored. Based on the literature reviewed here, microglia are a likely source of dysregulated miRNAs and potential mediators of neurodegenerative processes. Therefore, dysregulated miRNAs may be promising candidates for the development of therapeutic strategies.


Asunto(s)
Esclerosis Amiotrófica Lateral , Comunicación Celular/fisiología , MicroARNs/metabolismo , Microglía/metabolismo , Degeneración Nerviosa/metabolismo , Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/patología , Animales , Vesículas Extracelulares/metabolismo , Humanos , Microglía/patología , Degeneración Nerviosa/patología
3.
Neurobiol Dis ; 105: 283-299, 2017 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-28235672

RESUMEN

Intracellular trafficking of cargoes is an essential process to maintain the structure and function of all mammalian cell types, but especially of neurons because of their extreme axon/dendrite polarisation. Axonal transport mediates the movement of cargoes such as proteins, mRNA, lipids, membrane-bound vesicles and organelles that are mostly synthesised in the cell body and in doing so is responsible for their correct spatiotemporal distribution in the axon, for example at specialised sites such as nodes of Ranvier and synaptic terminals. In addition, axonal transport maintains the essential long-distance communication between the cell body and synaptic terminals that allows neurons to react to their surroundings via trafficking of for example signalling endosomes. Axonal transport defects are a common observation in a variety of neurodegenerative diseases, and mutations in components of the axonal transport machinery have unequivocally shown that impaired axonal transport can cause neurodegeneration (reviewed in El-Kadi et al., 2007, De Vos et al., 2008; Millecamps and Julien, 2013). Here we review our current understanding of axonal transport defects and the role they play in motor neuron diseases (MNDs) with a specific focus on the most common form of MND, amyotrophic lateral sclerosis (ALS).


Asunto(s)
Transporte Axonal/fisiología , Enfermedad de la Neurona Motora/fisiopatología , Neurobiología , Investigación Biomédica Traslacional/métodos , Animales , Humanos
4.
Hum Mol Genet ; 24(3): 828-40, 2015 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-25274775

RESUMEN

Aprataxin (APTX) deficiency causes progressive cerebellar degeneration, ataxia and oculomotor apraxia in man. Cell free assays and crystal structure studies demonstrate a role for APTX in resolving 5'-adenylated nucleic acid breaks, however, APTX function in vertebrates remains unclear due to the lack of an appropriate model system. Here, we generated a murine model in which a pathogenic mutant of superoxide dismutase 1 (SOD1(G93A)) is expressed in an Aptx-/- mouse strain. We report a delayed population doubling and accelerated senescence in Aptx-/- primary mouse fibroblasts, which is not due to detectable telomere instability or cell cycle deregulation but is associated with a reduction in transcription recovery following oxidative stress. Expression of SOD1(G93A) uncovers a survival defect ex vivo in cultured cells and in vivo in tissues lacking Aptx. The surviving neurons feature numerous and deep nuclear envelope invaginations, a hallmark of cellular stress. Furthermore, they possess an elevated number of high-density nuclear regions and a concomitant increase in histone H3 K9 trimethylation, hallmarks of silenced chromatin. Finally, the accelerated cellular senescence was also observed at the organismal level as shown by down-regulation of insulin-like growth factor 1 (IGF-1), a hallmark of premature ageing. Together, this study demonstrates a protective role of Aptx in vivo and suggests that its loss results in progressive accumulation of DNA breaks in the nervous system, triggering hallmarks of premature ageing, systemically.


Asunto(s)
Envejecimiento Prematuro/metabolismo , Proteínas de Unión al ADN/deficiencia , Neuronas Motoras/patología , Proteínas Nucleares/deficiencia , Superóxido Dismutasa/genética , Transcripción Genética/efectos de los fármacos , Envejecimiento Prematuro/genética , Envejecimiento Prematuro/patología , Animales , Células Cultivadas , Senescencia Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Humanos , Peróxido de Hidrógeno/farmacología , Factor I del Crecimiento Similar a la Insulina/metabolismo , Ratones , Mutación , Estrés Oxidativo , Superóxido Dismutasa/metabolismo , Superóxido Dismutasa-1
5.
Am J Hum Genet ; 92(6): 965-73, 2013 Jun 06.
Artículo en Inglés | MEDLINE | ID: mdl-23664120

RESUMEN

Dominant congenital spinal muscular atrophy (DCSMA) is a disorder of developing anterior horn cells and shows lower-limb predominance and clinical overlap with hereditary spastic paraplegia (HSP), a lower-limb-predominant disorder of corticospinal motor neurons. We have identified four mutations in bicaudal D homolog 2 (Drosophila) (BICD2) in six kindreds affected by DCSMA, DCSMA with upper motor neuron features, or HSP. BICD2 encodes BICD2, a key adaptor protein that interacts with the dynein-dynactin motor complex, which facilitates trafficking of cellular cargos that are critical to motor neuron development and maintenance. We demonstrate that mutations resulting in amino acid substitutions in two binding regions of BICD2 increase its binding affinity for the cytoplasmic dynein-dynactin complex, which might result in the perturbation of BICD2-dynein-dynactin-mediated trafficking, and impair neurite outgrowth. These findings provide insight into the mechanism underlying both the static and the slowly progressive clinical features and the motor neuron pathology that characterize BICD2-associated diseases, and underscore the importance of the dynein-dynactin transport pathway in the development and survival of both lower and upper motor neurons.


Asunto(s)
Proteínas Portadoras/genética , Atrofia Muscular Espinal/genética , Mutación Missense , Paraplejía/genética , Adulto , Anciano , Proteínas Portadoras/metabolismo , Niño , Preescolar , Dineínas Citoplasmáticas/metabolismo , Femenino , Genes Dominantes , Ligamiento Genético , Estudio de Asociación del Genoma Completo , Células HEK293 , Haplotipos , Humanos , Masculino , Proteínas Asociadas a Microtúbulos , Persona de Mediana Edad , Atrofia Muscular Espinal/congénito , Atrofia Muscular Espinal/metabolismo , Paraplejía/metabolismo , Linaje , Polimorfismo de Nucleótido Simple , Unión Proteica , Adulto Joven
6.
Nucleic Acids Res ; 42(1): 307-14, 2014 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-24049082

RESUMEN

Amyotrophic lateral sclerosis (ALS) is associated with progressive degeneration of motor neurons. Several of the genes associated with this disease encode proteins involved in RNA processing, including fused-in-sarcoma/translocated-in-sarcoma (FUS/TLS). FUS is a member of the heterogeneous nuclear ribonucleoprotein (hnRNP) family of proteins that bind thousands of pre-mRNAs and can regulate their splicing. Here, we have examined the possibility that FUS is also a component of the cellular response to DNA damage. We show that both GFP-tagged and endogenous FUS re-localize to sites of oxidative DNA damage induced by UVA laser, and that FUS recruitment is greatly reduced or ablated by an inhibitor of poly (ADP-ribose) polymerase activity. Consistent with this, we show that recombinant FUS binds directly to poly (ADP-ribose) in vitro, and that both GFP-tagged and endogenous FUS fail to accumulate at sites of UVA laser induced damage in cells lacking poly (ADP-ribose) polymerase-1. Finally, we show that GFP-FUS(R521G), harbouring a mutation that is associated with ALS, exhibits reduced ability to accumulate at sites of UVA laser-induced DNA damage. Together, these data suggest that FUS is a component of the cellular response to DNA damage, and that defects in this response may contribute to ALS.


Asunto(s)
Esclerosis Amiotrófica Lateral/genética , Daño del ADN , Poli(ADP-Ribosa) Polimerasas/fisiología , Proteína FUS de Unión a ARN/metabolismo , Animales , Células Cultivadas , Humanos , Ratones , Mutación , Oxidación-Reducción , Poli(ADP-Ribosa) Polimerasa-1 , Poli Adenosina Difosfato Ribosa/biosíntesis , Poli Adenosina Difosfato Ribosa/metabolismo , Proteína FUS de Unión a ARN/genética
7.
Brain ; 137(Pt 7): 1883-93, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24755273

RESUMEN

Mutations in the gene encoding the heavy chain subunit (DYNC1H1) of cytoplasmic dynein cause spinal muscular atrophy with lower extremity predominance, Charcot-Marie-Tooth disease and intellectual disability. We used the legs at odd angles (Loa) (DYNC1H1(F580Y)) mouse model for spinal muscular atrophy with lower extremity predominance and a combination of live-cell imaging and biochemical assays to show that the velocity of dynein-dependent microtubule minus-end (towards the nucleus) movement of EGF and BDNF induced signalling endosomes is significantly reduced in Loa embryonic fibroblasts and motor neurons. At the same time, the number of the plus-end (towards the cell periphery) moving endosomes is increased in the mutant cells. As a result, the extracellular signal-regulated kinases (ERK) 1/2 activation and c-Fos expression are altered in both mutant cell types, but the motor neurons exhibit a strikingly abnormal ERK1/2 and c-Fos response to serum-starvation induced stress. These data highlight the cell-type specific ERK1/2 response as a possible contributory factor in the neuropathological nature of Dync1h1 mutations, despite generic aberrant kinetics in both cell types, providing an explanation for how mutations in the ubiquitously expressed DYNC1H1 cause neuron-specific disease.


Asunto(s)
Dineínas Citoplasmáticas/genética , Sistema de Señalización de MAP Quinasas/genética , Atrofia Muscular Espinal/genética , Mutación/genética , Proteínas Proto-Oncogénicas c-fos/metabolismo , Animales , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Células Cultivadas , Medio de Cultivo Libre de Suero/farmacología , Modelos Animales de Enfermedad , Embrión de Mamíferos , Endosomas/efectos de los fármacos , Endosomas/metabolismo , Factor de Crecimiento Epidérmico/metabolismo , Humanos , Ratones , Ratones Transgénicos , Neuronas Motoras/efectos de los fármacos , Neuronas Motoras/metabolismo , Fosfoproteínas Fosfatasas/metabolismo , Transporte de Proteínas/efectos de los fármacos , Transporte de Proteínas/genética , Transfección
8.
J Cell Sci ; 125(Pt 16): 3733-8, 2012 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-22553211

RESUMEN

In migrating NRK cells, aPKCs control the dynamics of turnover of paxillin-containing focal adhesions (FA) determining migration rate. Using a proteomic approach (two-dimensional fluorescence difference gel electrophoresis), dynein intermediate chain 2 (dynein IC2) was identified as a protein that is phosphorylated inducibly during cell migration in a PKC-regulated manner. By gene silencing and co-immunoprecipitation studies, we show that dynein IC2 regulates the speed of cell migration through its interaction with paxillin. This interaction is controlled by serine 84 phosphorylation, which lies on the aPKC pathway. The evidence presented thus links aPKC control of migration to the dynein control of FA turnover through paxillin.


Asunto(s)
Adhesión Celular/fisiología , Movimiento Celular/fisiología , Dineínas/metabolismo , Paxillin/metabolismo , Secuencia de Aminoácidos , Animales , Células Cultivadas , Dineínas Citoplasmáticas , Riñón/citología , Riñón/enzimología , Datos de Secuencia Molecular , Fosforilación , Ratas
9.
Dis Model Mech ; 17(5)2024 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-38813848

RESUMEN

Evidence suggests the presence of microglial activation and microRNA (miRNA) dysregulation in amyotrophic lateral sclerosis (ALS), the most common form of adult motor neuron disease. However, few studies have investigated whether the miRNA dysregulation originates from microglia. Furthermore, TDP-43 (encoded by TARDBP), involved in miRNA biogenesis, aggregates in tissues of ∼98% of ALS cases. Thus, this study aimed to determine whether expression of the ALS-linked TDP-43M337V mutation in a transgenic mouse model dysregulates microglia-derived miRNAs. RNA sequencing identified several dysregulated miRNAs released by transgenic microglia and a differential miRNA release by lipopolysaccharide-stimulated microglia, which was more pronounced in cells from female mice. We validated the downregulation of three candidate miRNAs, namely, miR-16-5p, miR-99a-5p and miR-191-5p, by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and identified their predicted targets, which primarily include genes involved in neuronal development and function. These results suggest that altered TDP-43 function leads to changes in the miRNA population released by microglia, which may in turn be a source of the miRNA dysregulation observed in the disease. This has important implications for the role of neuroinflammation in ALS pathology and could provide potential therapeutic targets.


Asunto(s)
Esclerosis Amiotrófica Lateral , Ratones Transgénicos , MicroARNs , Microglía , Mutación , Caracteres Sexuales , Microglía/metabolismo , Microglía/patología , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/patología , MicroARNs/genética , MicroARNs/metabolismo , Animales , Femenino , Masculino , Mutación/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Ratones , Espacio Extracelular/metabolismo , Humanos , Lipopolisacáridos/farmacología , Regulación de la Expresión Génica
10.
Dis Model Mech ; 17(5)2024 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-38721655

RESUMEN

Evidence suggests the presence of microglial activation and microRNA (miRNA) dysregulation in amyotrophic lateral sclerosis (ALS), the most common form of adult motor neuron disease. However, few studies have investigated whether the miRNA dysregulation originates from microglia. Furthermore, TDP-43 (encoded by TARDBP), involved in miRNA biogenesis, aggregates in tissues of ∼98% of ALS cases. Thus, this study aimed to determine whether expression of the ALS-linked TDP-43M337V mutation in a transgenic mouse model dysregulates microglia-derived miRNAs. RNA sequencing identified several dysregulated miRNAs released by transgenic microglia and a differential miRNA release by lipopolysaccharide-stimulated microglia, which was more pronounced in cells from female mice. We validated the downregulation of three candidate miRNAs, namely, miR-16-5p, miR-99a-5p and miR-191-5p, by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and identified their predicted targets, which primarily include genes involved in neuronal development and function. These results suggest that altered TDP-43 function leads to changes in the miRNA population released by microglia, which may in turn be a source of the miRNA dysregulation observed in the disease. This has important implications for the role of neuroinflammation in ALS pathology and could provide potential therapeutic targets.


Asunto(s)
Esclerosis Amiotrófica Lateral , Ratones Transgénicos , MicroARNs , Microglía , Mutación , Caracteres Sexuales , Microglía/metabolismo , Microglía/patología , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/patología , MicroARNs/genética , MicroARNs/metabolismo , Animales , Femenino , Masculino , Mutación/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Ratones , Espacio Extracelular/metabolismo , Humanos , Lipopolisacáridos/farmacología , Regulación de la Expresión Génica
11.
Artículo en Inglés | MEDLINE | ID: mdl-37498094

RESUMEN

OBJECTIVE: Most TDP-43 mouse models of ALS do not display cytoplasmic mislocalisation or protein aggregation of TDP-43 in spinal motor neurons in vivo. Thus, we investigated whether a combination of defective dynein with a TDP-43 mutation could trigger TDP-43 pathology. METHODS: Using immunohistochemical methods we examined the intracellular motor neuron pathology of the offspring of TDP-43WT and TDP-43M337V transgenic mice bred to heterozygous Loa mice, which carry an autosomal dominant mutation in dynein cytoplasmic 1 heavy chain 1 (Dync1h1). RESULTS: These mice did not exhibit TDP-43 mislocalisation in spinal motor neurons, but the expression of mutant dynein in combination with wildtype human TDP-43 resulted in p62 upregulation and TDP-43 aggregation, thus partially recapitulating the human disease. CONCLUSIONS: These findings provide new insights into the possible relationship between dynein and TDP-43 and could prove useful in future studies looking to elucidate the mechanism behind the TDP-43 pathology observed in ALS.

12.
J Neurosci ; 31(14): 5483-94, 2011 Apr 06.
Artículo en Inglés | MEDLINE | ID: mdl-21471385

RESUMEN

The cytoplasmic dynein complex is fundamentally important to all eukaryotic cells for transporting a variety of essential cargoes along microtubules within the cell. This complex also plays more specialized roles in neurons. The complex consists of 11 types of protein that interact with each other and with external adaptors, regulators and cargoes. Despite the importance of the cytoplasmic dynein complex, we know comparatively little of the roles of each component protein, and in mammals few mutants exist that allow us to explore the effects of defects in dynein-controlled processes in the context of the whole organism. Here we have taken a genotype-driven approach in mouse (Mus musculus) to analyze the role of one subunit, the dynein light intermediate chain 1 (Dync1li1). We find that, surprisingly, an N235Y point mutation in this protein results in altered neuronal development, as shown from in vivo studies in the developing cortex, and analyses of electrophysiological function. Moreover, mutant mice display increased anxiety, thus linking dynein functions to a behavioral phenotype in mammals for the first time. These results demonstrate the important role that dynein-controlled processes play in the correct development and function of the mammalian nervous system.


Asunto(s)
Conducta Animal/fisiología , Dineínas Citoplasmáticas/genética , Regulación del Desarrollo de la Expresión Génica/genética , Fenotipo , Mutación Puntual/genética , Animales , Animales Recién Nacidos , Asparagina/genética , Recuento de Células/métodos , Células Cultivadas , Corteza Cerebral/citología , Dendritas/genética , Embrión de Mamíferos , Femenino , Fibroblastos/fisiología , Fibroblastos/ultraestructura , Ganglios Espinales/citología , Proteínas Fluorescentes Verdes/genética , Masculino , Aprendizaje por Laberinto/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Actividad Motora/genética , Proteínas del Tejido Nervioso , Conducción Nerviosa/genética , Neuronas/clasificación , Neuronas/citología , Neuronas/fisiología , Transporte de Proteínas/efectos de los fármacos , Transporte de Proteínas/genética , Desempeño Psicomotor , Estadísticas no Paramétricas , Tirosina/genética , Levantamiento de Peso/fisiología
13.
Biochim Biophys Acta ; 1812(1): 59-69, 2011 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-20887786

RESUMEN

The molecular motor dynein is regulated by the huntingtin protein, and Huntington's disease (HD) mutations of huntingtin disrupt dynein motor activity. Besides abnormalities in the central nervous system, HD animal models develop prominent peripheral pathology, with defective brown tissue thermogenesis and dysfunctional white adipocytes, but whether this peripheral phenotype is recapitulated by dynein dysfunction is unknown. Here, we observed prominently increased adiposity in mice harboring the legs at odd angles (Loa/+) or the Cramping mutations (Cra/+) in the dynein heavy chain gene. In Cra/+ mice, hyperadiposity occurred in the absence of energy imbalance and was the result of impaired norepinephrine-stimulated lipolysis. A similar phenotype was observed in 3T3L1 adipocytes upon chemical inhibition of dynein showing that loss of functional dynein leads to impairment of lipolysis. Ex vivo, dynein mutant adipose tissue displayed increased reactive oxygen species production that was, at least partially, responsible for the decreased cellular responses to norepinephrine and subsequent defect in stimulated lipolysis. Dynein mutation also affected norepinephrine efficacy to elicit a thermogenic response and led to morphological abnormalities in brown adipose tissue and cold intolerance in dynein mutant mice. Interestingly, protein levels of huntingtin were decreased in dynein mutant adipose tissue. Collectively, our results provide genetic evidence that dynein plays a key role in lipid metabolism and thermogenesis through a modulation of oxidative stress elicited by norepinephrine. This peripheral phenotype of dynein mutant mice is similar to that observed in various animal models of HD, lending further support for a functional link between huntingtin and dynein.


Asunto(s)
Tejido Adiposo Pardo/metabolismo , Tejido Adiposo Blanco/metabolismo , Dineínas Citoplasmáticas/genética , Metabolismo Energético/genética , Mutación , Células 3T3-L1 , Agonistas alfa-Adrenérgicos/farmacología , Animales , Western Blotting , Dineínas Citoplasmáticas/metabolismo , Femenino , Expresión Génica , Humanos , Proteína Huntingtina , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Lipólisis/efectos de los fármacos , Lipólisis/genética , Masculino , Ratones , Ratones Mutantes , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Norepinefrina/farmacología , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Estrés Oxidativo/efectos de los fármacos , Receptores Adrenérgicos beta 2/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal , Termogénesis/genética
14.
Bull Math Biol ; 74(9): 2032-61, 2012 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-22791512

RESUMEN

Mutations in the motor protein cytoplasmic dynein have been found to cause Charcot-Marie-Tooth disease, spinal muscular atrophy, and severe intellectual disabilities in humans. In mouse models, neurodegeneration is observed. We sought to develop a novel model which could incorporate the effects of mutations on distance travelled and velocity. A mechanical model for the dynein mediated transport of endosomes is derived from first principles and solved numerically. The effects of variations in model parameter values are analysed to find those that have a significant impact on velocity and distance travelled. The model successfully describes the processivity of dynein and matches qualitatively the velocity profiles observed in experiments.


Asunto(s)
Membrana Celular/metabolismo , Núcleo Celular/metabolismo , Dineínas Citoplasmáticas/metabolismo , Modelos Biológicos , Adenosina Trifosfato/metabolismo , Transporte Biológico , Endocitosis , Receptores ErbB/metabolismo , Análisis Numérico Asistido por Computador
15.
Ann Med ; 54(1): 3069-3078, 2022 12.
Artículo en Inglés | MEDLINE | ID: mdl-36314539

RESUMEN

INTRODUCTION: Objective biomarkers for the fatal neurodegenerative disease amyotrophic lateral sclerosis or motor neuron disease (ALS/MND) are critical for diagnosis, drug development, clinical trials, and insight into disease pathology. Key candidates for biomarkers present in biofluids include non-coding RNA (ncRNA) transcripts including microRNA, piwi-interacting RNA and transfer RNA. To determine if the central nervous system was the source of the dysregulated ncRNA biomarkers we previously observed in serum, we sought to identify dysregulated ncRNA candidates in cerebrospinal fluid (CSF) which may provide new insight into the disease pathology. METHODS AND MATERIALS: Small RNA sequencing (RNA-seq) was undertaken on CSF samples from healthy controls (n = 18), disease mimics (n = 8), and ALS patients (n = 40) in our Oxford Study for Biomarkers of ALS cohort, with RT-qPCR used to confirm their dysregulation. RESULTS: We found a range of ncRNA that were dysregulated in the RNA-seq screen, but these failed to be validated or detected in some cases using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Additionally, our previously identified serum ncRNA biomarker showed no change in CSF or correlation to serum. CONCLUSIONS: This study suggests the CSF may not be the source of dysregulated ncRNA in the serum and highlights the difficulty in identifying ncRNA in CSF as biomarkers for ALS.KEY MESSAGESIn this current study, we investigated the expression of non-coding RNA transcripts in the cerebrospinal fluid of ALS patients compared to healthy controls.RNA-seq identified dysregulated non-coding RNA transcripts, but these were not validated with RT-qPCR.We conclude that cerebrospinal fluid is not a suitable source of diagnostic biomarkers.


Asunto(s)
Esclerosis Amiotrófica Lateral , MicroARNs , Enfermedades Neurodegenerativas , Humanos , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/diagnóstico , Esclerosis Amiotrófica Lateral/patología , Biomarcadores , Estudios de Cohortes
16.
J Biol Chem ; 285(51): 39922-34, 2010 Dec 17.
Artículo en Inglés | MEDLINE | ID: mdl-20889981

RESUMEN

A single amino acid change, F580Y (Legs at odd angles (Loa), Dync1h1(Loa)), in the highly conserved and overlapping homodimerization, intermediate chain, and light intermediate chain binding domain of the cytoplasmic dynein heavy chain can cause severe motor and sensory neuron loss in mice. The mechanism by which the Loa mutation impairs the neuron-specific functions of dynein is not understood. To elucidate the underlying molecular mechanisms of neurodegeneration arising from this mutation, we applied a cohort of biochemical methods combined with in vivo assays to systemically study the effects of the mutation on the assembly of dynein and its interaction with dynactin. We found that the Loa mutation in the heavy chain leads to increased affinity of this subunit of cytoplasmic dynein to light intermediate and a population of intermediate chains and a suppressed association of dynactin to dynein. These data suggest that the Loa mutation drives the assembly of cytoplasmic dynein toward a complex with lower affinity to dynactin and thus impairing transport of cargos that tether to the complex via dynactin. In addition, we detected up-regulation of kinesin light chain 1 (KLC1) and its increased association with dynein but reduced microtubule-associated KLC1 in the Loa samples. We provide a model describing how up-regulation of KLC1 and its interaction with cytoplasmic dynein in Loa could play a regulatory role in restoring the retrograde and anterograde transport in the Loa neurons.


Asunto(s)
Dineínas Citoplasmáticas/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Mutación , Enfermedades Neurodegenerativas/metabolismo , Animales , Dineínas Citoplasmáticas/genética , Complejo Dinactina , Cinesinas , Ratones , Ratones Mutantes , Proteínas Asociadas a Microtúbulos/genética , Enfermedades Neurodegenerativas/genética
17.
J Biol Chem ; 285(24): 18627-39, 2010 Jun 11.
Artículo en Inglés | MEDLINE | ID: mdl-20382740

RESUMEN

Amyotrophic lateral sclerosis (ALS) is a debilitating and fatal late-onset neurodegenerative disease. Familial cases of ALS (FALS) constitute approximately 10% of all ALS cases, and mutant superoxide dismutase 1 (SOD1) is found in 15-20% of FALS. SOD1 mutations confer a toxic gain of unknown function to the protein that specifically targets the motor neurons in the cortex and the spinal cord. We have previously shown that the autosomal dominant Legs at odd angles (Loa) mutation in cytoplasmic dynein heavy chain (Dync1h1) delays disease onset and extends the life span of transgenic mice harboring human mutant SOD1(G93A). In this study we provide evidence that despite the lack of direct interactions between mutant SOD1 and either mutant or wild-type cytoplasmic dynein, the Loa mutation confers significant reductions in the amount of mutant SOD1 protein in the mitochondrial matrix. Moreover, we show that the Loa mutation ameliorates defects in mitochondrial respiration and membrane potential observed in SOD1(G93A) motor neuron mitochondria. These data suggest that the Loa mutation reduces the vulnerability of mitochondria to the toxic effects of mutant SOD1, leading to improved mitochondrial function in SOD1(G93A) motor neurons.


Asunto(s)
Modelos Animales de Enfermedad , Dineínas/genética , Mitocondrias/metabolismo , Enfermedad de la Neurona Motora/metabolismo , Mutación , Superóxido Dismutasa/genética , Animales , Citoplasma/metabolismo , Femenino , Heterocigoto , Potenciales de la Membrana , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Neuronas Motoras/metabolismo , Superóxido Dismutasa-1
18.
J Cell Biol ; 169(4): 561-7, 2005 May 23.
Artículo en Inglés | MEDLINE | ID: mdl-15911875

RESUMEN

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative condition characterized by motoneuron degeneration and muscle paralysis. Although the precise pathogenesis of ALS remains unclear, mutations in Cu/Zn superoxide dismutase (SOD1) account for approximately 20-25% of familial ALS cases, and transgenic mice overexpressing human mutant SOD1 develop an ALS-like phenotype. Evidence suggests that defects in axonal transport play an important role in neurodegeneration. In Legs at odd angles (Loa) mice, mutations in the motor protein dynein are associated with axonal transport defects and motoneuron degeneration. Here, we show that retrograde axonal transport defects are already present in motoneurons of SOD1(G93A) mice during embryonic development. Surprisingly, crossing SOD1(G93A) mice with Loa/+ mice delays disease progression and significantly increases life span in Loa/SOD1(G93A) mice. Moreover, there is a complete recovery in axonal transport deficits in motoneurons of these mice, which may be responsible for the amelioration of disease. We propose that impaired axonal transport is a prime cause of neuronal death in neurodegenerative disorders such as ALS.


Asunto(s)
Esclerosis Amiotrófica Lateral/metabolismo , Transporte Axonal/genética , Dineínas/genética , Mutación/genética , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/fisiopatología , Animales , Axones/metabolismo , Axones/patología , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Dineínas/biosíntesis , Femenino , Humanos , Masculino , Ratones , Ratones Mutantes Neurológicos , Ratones Transgénicos , Neuronas Motoras/metabolismo , Neuronas Motoras/patología , Degeneración Nerviosa/genética , Degeneración Nerviosa/metabolismo , Degeneración Nerviosa/fisiopatología , Recuperación de la Función/genética , Superóxido Dismutasa/genética , Tasa de Supervivencia
19.
Brain Commun ; 2(1): fcaa053, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32613197

RESUMEN

Objective biomarkers for the clinically heterogeneous adult-onset neurodegenerative disorder amyotrophic lateral sclerosis are crucial to facilitate assessing emerging therapeutics and improve the diagnostic pathway in what is a clinically heterogeneous syndrome. With non-coding RNA transcripts including microRNA, piwi-RNA and transfer RNA present in human biofluids, we sought to identify whether non-coding RNA in serum could be biomarkers for amyotrophic lateral sclerosis. Serum samples from our Oxford Study for Biomarkers in motor neurone disease/amyotrophic lateral sclerosis discovery cohort of amyotrophic lateral sclerosis patients (n = 48), disease mimics (n = 16) and age- and sex-matched healthy controls (n = 24) were profiled for non-coding RNA expression using RNA-sequencing, which showed a wide range of non-coding RNA to be dysregulated. We confirmed significant alterations with reverse transcription-quantitative PCR in the expression of hsa-miR-16-5p, hsa-miR-21-5p, hsa-miR-92a-3p, hsa-piR-33151, TRV-AAC4-1.1 and TRA-AGC6-1.1. Furthermore, hsa-miR-206, a previously identified amyotrophic lateral sclerosis biomarker, showed a binary-like pattern of expression in our samples. Using the expression of these non-coding RNA, we were able to discriminate amyotrophic lateral sclerosis samples from healthy controls in our discovery cohort using a random forest analysis with 93.7% accuracy with promise in predicting progression rate of patients. Importantly, cross-validation of this novel signature using a new geographically distinct cohort of samples from the United Kingdom and Germany with both amyotrophic lateral sclerosis and control samples (n = 156) yielded an accuracy of 73.9%. The high prediction accuracy of this non-coding RNA-based biomarker signature, even across heterogeneous cohorts, demonstrates the strength of our approach as a novel platform to identify and stratify amyotrophic lateral sclerosis patients.

20.
Front Neurol ; 10: 186, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-30899244

RESUMEN

Amyotrophic lateral sclerosis (ALS; MND, motor neuron disease) is a debilitating neurodegenerative disease affecting 4.5 per 100,000 people per year around the world. There is currently no cure for this disease, and its causes are relatively unknown. Diagnosis is based on a battery of clinical tests up to a year after symptom onset, with no robust markers of diagnosis or disease progression currently identified. A major thrust of current research is to identify potential non-invasive markers ("biomarkers") in body fluids such as blood and/or cerebrospinal fluid (CSF) to use for diagnostic or prognostic purposes. Non-coding RNAs (ncRNAs), including microRNAs (miRNAs), are found at detectable and stable levels in blood and other bodily fluids. Specific ncRNAs can vary in levels between ALS patients and non-ALS controls without the disease. In this review, we will provide an overview of early findings, demonstrate the potential of this new class as biomarkers, and discuss future challenges and opportunities taking this forward to help patients with ALS.

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