RESUMEN
BACKGROUND: Mutations in several genes of Caenorhabditis elegans confer altered sensitivities to volatile anesthetics. A mutation in one gene, gas-1(fc21), causes animals to be immobilized at lower concentrations of all volatile anesthetics than in the wild type, and it does not depend on mutations in other genes to control anesthetic sensitivity. gas-1 confers different sensitivities to stereoisomers of isoflurane, and thus may be a direct target for volatile anesthetics. The authors have cloned and characterized the gas gene and the mutant allele fc21. METHODS: Genetic techniques for nematodes were as previously described. Polymerase chain reaction, sequencing, and other molecular biology techniques were performed by standard methods. Mutant rescue was done by injecting DNA fragments into the gonad of mutant animals and scoring the offspring for loss of the mutant phenotype. RESULTS: The gas-1 gene was cloned and identified. The protein GAS-1 is a homologue of the 49-kd (IP) subunit of the mitochondrial NADH-ubiquinone-oxidoreductase (complex I of the respiratory chain). gas-1(fc21) is a missense mutation replacing a strictly conserved arginine with lysine. CONCLUSIONS: The function of the 49-kd (IP) subunit of complex I is unknown. The finding that mutations in complex I increase sensitivity of C. elegans to volatile anesthetics may implicate this physiologic process in the determination of anesthetic sensitivity. The hypersensitivity of animals with a mutation in the gas-1 gene may be caused by a direct anesthetic effect on a mitochondrial protein or secondary effects at other sites caused by mitochondrial dysfunction.
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Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animales , Caenorhabditis elegans/genética , Caenorhabditis elegans/efectos de los fármacos , Proteínas de Caenorhabditis elegans/genética , Mutación , Anestésicos por Inhalación , Complejo I de Transporte de Electrón/genética , Complejo I de Transporte de Electrón/metabolismo , Secuencia de Aminoácidos , Isoflurano/farmacologíaRESUMEN
Panax ginseng C. A. Meyer (ginseng) is a medicinal plant widely used for promoting longevity. Recently, homogalacturonan (HG) domain-rich pectins purified from some plants have been reported to have anti-aging-related activities, leading us to explore the longevity-promoting activity of the HG pectins from ginseng. In this study, we discovered that two of low methyl-esterified ginseng HG pectins (named as WGPA-2-HG and WGPA-3-HG), whose degree of methyl-esterification (DM) was 16 % and 8 % respectively, promoted longevity in Caenorhabditis elegans. Results showed that WGPA-2-HG/WGPA-3-HG impaired insulin/insulin-like growth factor 1 (IGF-1) signalling (IIS) pathway, thereby increasing the nuclear accumulation of transcription factors SKN-1/Nrf2 and DAF-16/FOXO and enhancing the expression of relevant anti-aging genes. BLI and ITC analysis showed that the insulin-receptor binding, the first step to activate IIS pathway, was impeded by the engagement of WGPA-2-HG/WGPA-3-HG with insulin. By chemical modifications, we found that high methyl-esterification of WGPA-2-HG/WGPA-3-HG was detrimental for their longevity-promoting activity. These findings provided novel insight into the precise molecular mechanism for the longevity-promoting effect of ginseng pectins, and suggested a potential to utilize the ginseng HG pectins with appropriate DM values as natural nutrients for increasing human longevity.
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Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Factor I del Crecimiento Similar a la Insulina , Insulina , Longevidad , Panax , Pectinas , Transducción de Señal , Animales , Caenorhabditis elegans/efectos de los fármacos , Caenorhabditis elegans/metabolismo , Panax/química , Factor I del Crecimiento Similar a la Insulina/metabolismo , Pectinas/farmacología , Pectinas/metabolismo , Pectinas/química , Longevidad/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Insulina/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , EsterificaciónAsunto(s)
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animales , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Acoplamiento Excitación-Contracción/fisiología , Células Musculares/metabolismo , Canales de Potasio de Gran Conductancia Activados por el Calcio/metabolismoRESUMEN
Measuring neuronal activity is important for understanding neuronal function. Ca2+ imaging by genetically encoded calcium indicators (GECIs) is a powerful way to measure neuronal activity. Although it revealed important aspects of neuronal function, measuring the neuronal membrane voltage is important to understand neuronal function as it triggers neuronal activation. Recent progress of genetically encoded voltage indicators (GEVIs) enabled us fast and precise measurements of neuronal membrane voltage. To clarify the relation of the membrane voltage and intracellular Ca2+, we analyzed neuronal activities of olfactory neuron AWA in Caenorhabditis elegans by GCaMP6f (GECI) and paQuasAr3 (GEVI) responding to odorants. We found that the membrane voltage encodes the stimuli change by the timing and the duration by the weak semi-stable depolarization. However, the change of the intracellular Ca2+ encodes the strength of the stimuli. Furthermore, ODR-3, a G-protein alpha subunit, was shown to be important for stabilizing the membrane voltage. These results suggest that the combination of calcium and voltage imaging provides a deeper understanding of the information in neural circuits.
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Caenorhabditis elegans , Calcio , Animales , Caenorhabditis elegans/fisiología , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/genética , Calcio/metabolismo , Potenciales de la Membrana/fisiología , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Neuronas Receptoras Olfatorias/fisiología , Neuronas Receptoras Olfatorias/metabolismo , Odorantes/análisisRESUMEN
When an organism encounters a pathogen, the host innate immune system activates to defend against pathogen colonization and toxic xenobiotics produced. C. elegans employ multiple defense systems to ensure survival when exposed to Pseudomonas aeruginosa including activation of the cytoprotective transcription factor SKN-1/NRF2. Although wildtype C. elegans quickly learn to avoid pathogens, here we describe a peculiar apathy-like behavior towards PA14 in animals with constitutive activation of SKN-1, whereby animals choose not to leave and continue to feed on the pathogen even when a non-pathogenic and healthspan-promoting food option is available. Although lacking the urgency to escape the infectious environment, animals with constitutive SKN-1 activity are not oblivious to the presence of the pathogen and display the typical pathogen-induced intestinal distension and eventual demise. SKN-1 activation, specifically in neurons and intestinal tissues, orchestrates a unique transcriptional program which leads to defects in serotonin signaling that is required from both neurons and non-neuronal tissues. Serotonin depletion from SKN-1 activation limits pathogen defenses capacity, drives the pathogen-associated apathy behaviors and induces a synthetic sensitivity to selective serotonin reuptake inhibitors. Taken together, our work reveals interesting insights into how animals perceive environmental pathogens and subsequently alter behavior and cellular programs to promote survival.
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Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Proteínas de Unión al ADN , Pseudomonas aeruginosa , Serotonina , Factores de Transcripción , Animales , Caenorhabditis elegans/microbiología , Caenorhabditis elegans/inmunología , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Pseudomonas aeruginosa/fisiología , Pseudomonas aeruginosa/patogenicidad , Serotonina/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteínas de Unión al ADN/genética , Neuronas/metabolismo , Infecciones por Pseudomonas/inmunología , Infecciones por Pseudomonas/metabolismo , Infecciones por Pseudomonas/microbiología , Inmunidad Innata , Transducción de Señal , Apatía/fisiología , Interacciones Huésped-Patógeno/inmunología , Inhibidores Selectivos de la Recaptación de Serotonina/farmacologíaRESUMEN
How can inter-individual variability be quantified? Measuring many features per experiment raises the question of choosing them to recapitulate high-dimensional data. Tackling this challenge on spindle elongation phenotypes, we showed that only three typical elongation patterns describe spindle elongation in C. elegans one-cell embryo. These archetypes, automatically extracted from the experimental data using principal component analysis (PCA), accounted for more than 95% of inter-individual variability of more than 1600 experiments across more than 100 different conditions. The two first archetypes were related to spindle average length and anaphasic elongation rate. The third archetype, accounting for 6% of the variability, was novel and corresponded to a transient spindle shortening in late metaphase, reminiscent of kinetochore function-defect phenotypes. Importantly, these three archetypes were robust to the choice of the dataset and were found even considering only non-treated conditions. Thus, the inter-individual differences between genetically perturbed embryos have the same underlying nature as natural inter-individual differences between wild-type embryos, independently of the temperatures. We thus propose that beyond the apparent complexity of the spindle, only three independent mechanisms account for spindle elongation, weighted differently in the various conditions. Interestingly, the spindle-length archetypes covered both metaphase and anaphase, suggesting that spindle elongation in late metaphase is sufficient to predict the late anaphase length. We validated this idea using a machine-learning approach. Finally, given amounts of these three archetypes could represent a quantitative phenotype. To take advantage of this, we set out to predict interacting genes from a seed based on the PCA coefficients. We exemplified this firstly on the role of tpxl-1 whose homolog tpx2 is involved in spindle microtubule branching, secondly the mechanism regulating metaphase length, and thirdly the central spindle players which set the length at anaphase. We found novel interactors not in public databases but supported by recent experimental publications.
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Caenorhabditis elegans , Fenotipo , Huso Acromático , Caenorhabditis elegans/embriología , Caenorhabditis elegans/fisiología , Caenorhabditis elegans/genética , Huso Acromático/fisiología , Animales , Análisis de Componente Principal , Biología Computacional , Embrión no Mamífero/embriología , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismoRESUMEN
The development of functional neurons is a complex orchestration of multiple signaling pathways controlling cell proliferation and differentiation. Because the balance of antioxidants is important for neuronal survival and development, we hypothesized that ferroptosis must be suppressed to gain neurons. We find that removal of antioxidants diminishes neuronal development and laminar organization of cortical organoids, which is fully restored when ferroptosis is inhibited by ferrostatin-1 or when neuronal differentiation occurs in the presence of vitamin A. Furthermore, iron-overload-induced developmental growth defects in C. elegans are ameliorated by vitamin E and A. We determine that all-trans retinoic acid activates the Retinoic Acid Receptor, which orchestrates the expression of anti-ferroptotic genes. In contrast, retinal and retinol show radical-trapping antioxidant activity. Together, our study reveals an unexpected function of vitamin A in coordinating the expression of essential cellular gatekeepers of ferroptosis, and demonstrates that suppression of ferroptosis by radical-trapping antioxidants or by vitamin A is required to obtain mature neurons and proper laminar organization in cortical organoids.
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Antioxidantes , Caenorhabditis elegans , Ferroptosis , Neuronas , Vitamina A , Animales , Ferroptosis/efectos de los fármacos , Vitamina A/farmacología , Vitamina A/metabolismo , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/efectos de los fármacos , Antioxidantes/farmacología , Neuronas/metabolismo , Neuronas/efectos de los fármacos , Neuronas/citología , Ciclohexilaminas/farmacología , Diferenciación Celular/efectos de los fármacos , Vitamina E/farmacología , Receptores de Ácido Retinoico/metabolismo , Receptores de Ácido Retinoico/genética , Tretinoina/farmacología , Organoides/efectos de los fármacos , Organoides/metabolismo , Neurogénesis/efectos de los fármacos , Ratones , Humanos , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Transducción de Señal/efectos de los fármacos , FenilendiaminasRESUMEN
Detecting chemical signals is important for identifying food sources and avoiding harmful agents. Like many animals, C. elegans use olfaction to chemotax towards their main food source, bacteria. However, little is known about the bacterial compounds governing C. elegans attraction to bacteria and the physiological importance of these compounds to bacteria. Here, we address these questions by investigating the function of a small RNA, P11, in the pathogen, Pseudomonas aeruginosa, that was previously shown to mediate learned pathogen avoidance. We discovered that this RNA also affects the attraction of untrained C. elegans to P. aeruginosa and does so by controlling production of ammonia, a volatile odorant produced during nitrogen assimilation. We describe the complex regulation of P. aeruginosa nitrogen assimilation, which is mediated by a partner-switching mechanism involving environmental nitrates, sensor proteins, and P11. In addition to mediating C. elegans attraction, we demonstrate that nitrogen assimilation mutants perturb bacterial fitness and pathogenesis during C. elegans infection by P. aeruginosa. These studies define ammonia as a major mediator of trans-kingdom signaling, implicate nitrogen assimilation as important for both bacteria and host organisms, and highlight how a bacterial metabolic pathway can either benefit or harm a host in different contexts.
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Amoníaco , Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Nitrógeno , Pseudomonas aeruginosa , Caenorhabditis elegans/microbiología , Caenorhabditis elegans/metabolismo , Pseudomonas aeruginosa/metabolismo , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/fisiología , Animales , Nitrógeno/metabolismo , Amoníaco/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/genética , Infecciones por Pseudomonas/microbiología , Infecciones por Pseudomonas/metabolismo , Nitratos/metabolismo , Transducción de Señal , Interacciones Huésped-Patógeno , QuimiotaxisRESUMEN
Clerodendranthus spicatus (Thunb.) (Kidney tea) is a very distinctive ethnic herbal medicine in China. Its leaves are widely used as a healthy tea. Many previous studies have demonstrated its various longevity-promoting effects; however, the safety and specific health-promoting effects of Clerodendranthus spicatus (C. spicatus) as a dietary supplement remain unclear. In order to understand the effect of C. spicatus on the longevity of Caenorhabditis elegans (C. elegans), we evaluated its role in C. elegans; C. spicatus water extracts (CSw) were analyzed for the major components and the effects on C. elegans were investigated from physiological and biochemical to molecular levels; CSw contain significant phenolic components (primarily rosmarinic acid and eugenolinic acid) and flavonoids (primarily quercetin and isorhamnetin) and can increase the lifespan of C. elegans. Further investigations showed that CSw modulate stress resistance and lipid metabolism through influencing DAF-16/FoxO (DAF-16), Heat shock factor 1 (HSF-1), and Nuclear Hormone Receptor-49 (NHR-49) signalling pathways; CSw can improve the antioxidant and hypolipidemic activity of C. elegans and prolong the lifespan of C. elegans (with the best effect at low concentrations). Therefore, the recommended daily use of C. spicatus should be considered when consuming it as a healthy tea on a daily basis.
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Caenorhabditis elegans , Metabolismo de los Lípidos , Estrés Oxidativo , Extractos Vegetales , Animales , Caenorhabditis elegans/efectos de los fármacos , Caenorhabditis elegans/metabolismo , Estrés Oxidativo/efectos de los fármacos , Extractos Vegetales/farmacología , Extractos Vegetales/química , Metabolismo de los Lípidos/efectos de los fármacos , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Longevidad/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Antioxidantes/farmacología , AguaRESUMEN
Glycation Stress (GS), induced by advanced glycation end-products (AGEs), significantly impacts aging processes. This study introduces a new model of GS of Caenorhabditis elegans by feeding them Escherichia coli OP50 cultured in a glucose-enriched medium, which better simulates human dietary glycation compared to previous single protein-glucose cross-linking methods. Utilizing WormCNN, a deep learning model, we assessed the health status and calculated the Healthy Aging Index (HAI) of worms with or without GS. Our results demonstrated accelerated aging in the GS group, evidenced by increased autofluorescence and altered gene expression of key aging regulators, daf-2 and daf-16. Additionally, we observed elevated pharyngeal pumping rates in AGEs-fed worms, suggesting an addictive response similar to human dietary patterns. This study highlights the profound effects of GS on worm aging and underscores the critical role of computer vision in accurately assessing health status and aiding in the establishment of disease models. The findings provide insights into glycation-induced aging and offer a comprehensive approach to studying the effects of dietary glycation on aging processes.
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Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Productos Finales de Glicación Avanzada , Animales , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/genética , Productos Finales de Glicación Avanzada/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Envejecimiento Saludable/metabolismo , Envejecimiento/metabolismo , Estrés Fisiológico , Factores de Transcripción Forkhead/metabolismo , Factores de Transcripción Forkhead/genética , Glicosilación , Glucosa/metabolismo , Modelos Animales de Enfermedad , Receptor de InsulinaRESUMEN
Although centrosomes help organize spindles in most cell types, oocytes of most species lack these structures. During acentrosomal spindle assembly in C. elegans oocytes, microtubule minus ends are sorted outwards away from the chromosomes where they form poles, but then these outward forces must be balanced to form a stable bipolar structure. Simultaneously, microtubule dynamics must be precisely controlled to maintain spindle length and organization. How forces and dynamics are tuned to create a stable bipolar structure is poorly understood. Here, we have gained insight into this question through studies of ZYG-8, a conserved doublecortin-family kinase; the mammalian homolog of this microtubule-associated protein is upregulated in many cancers and has been implicated in cell division, but the mechanisms by which it functions are poorly understood. We found that ZYG-8 depletion from oocytes resulted in overelongated spindles with pole and midspindle defects. Importantly, experiments with monopolar spindles revealed that ZYG-8 depletion led to excess outward forces within the spindle and suggested a potential role for this protein in regulating the force-generating motor BMK-1/kinesin-5. Further, we found that ZYG-8 is also required for proper microtubule dynamics within the oocyte spindle and that kinase activity is required for its function during both meiosis and mitosis. Altogether, our findings reveal new roles for ZYG-8 in oocytes and provide insights into how acentrosomal spindles are stabilized to promote faithful meiosis.
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Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Microtúbulos , Oocitos , Huso Acromático , Animales , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/fisiología , Microtúbulos/metabolismo , Microtúbulos/genética , Huso Acromático/metabolismo , Huso Acromático/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Oocitos/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Asociadas a Microtúbulos/genética , Centrosoma/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genéticaRESUMEN
The Caenorhabditis elegans HMP-2/HMP-1 complex, akin to the mammalian [Formula: see text]-catenin-[Formula: see text]-catenin complex, serves as a critical mechanosensor at cell-cell adherens junctions, transducing tension between HMR-1 (also known as cadherin in mammals) and the actin cytoskeleton. Essential for embryonic development and tissue integrity in C. elegans, this complex experiences tension from both internal actomyosin contractility and external mechanical microenvironmental perturbations. While offering a valuable evolutionary comparison to its mammalian counterpart, the impact of tension on the mechanical stability of HMP-1 and HMP-2/HMP-1 interactions remains unexplored. In this study, we directly quantified the mechanical stability of full-length HMP-1 and its force-bearing modulation domains (M1-M3), as well as the HMP-2/HMP-1 interface. Notably, the M1 domain in HMP-1 exhibits significantly higher mechanical stability than its mammalian analog, attributable to interdomain interactions with M2-M3. Introducing salt bridge mutations in the M3 domain weakens the mechanical stability of the M1 domain. Moreover, the intermolecular HMP-2/HMP-1 interface surpasses its mammalian counterpart in mechanical stability, enabling it to support the mechanical activation of the autoinhibited M1 domain for mechanotransduction. Additionally, the phosphomimetic mutation Y69E in HMP-2 weakens the mechanical stability of the HMP-2/HMP-1 interface, compromising the force-transmission molecular linkage and its associated mechanosensing functions. Collectively, these findings provide mechanobiological insights into the C. elegans HMP-2/HMP-1 complex, highlighting the impact of salt bridges on mechanical stability in [Formula: see text]-catenin and demonstrating the evolutionary conservation of the mechanical switch mechanism activating the HMP-1 modulation domain for protein binding at the single-molecule level.
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Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Mecanotransducción Celular , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/química , Proteínas de Caenorhabditis elegans/genética , Animales , Caenorhabditis elegans/metabolismo , Mecanotransducción Celular/fisiología , Imagen Individual de Molécula , Unión Proteica , Cadherinas/metabolismo , Cadherinas/química , Cadherinas/genética , Uniones Adherentes/metabolismo , Citoesqueleto de Actina/metabolismo , Citoesqueleto de Actina/química , Proteínas del Citoesqueleto , alfa CateninaRESUMEN
Mutations in the presenilin (PS) genes are a predominant cause of familial Alzheimer's disease (fAD). An ortholog of PS in the genetic model organism Caenorhabditis elegans (C. elegans) is sel-12. Mutations in the presenilin genes are commonly thought to lead to fAD by upregulating the expression of amyloid beta (Aß), however this hypothesis has been challenged by recent evidence. As C. elegans lack amyloid beta (Aß), the goal of this work was to examine Aß-independent effects of mutations in sel-12 and PS1/PS2 on behaviour and sensory neuron morphology across the lifespan in a C. elegans model. Olfactory chemotaxis experiments were conducted on sel-12(ok2078) loss-of-function mutant worms. Adult sel-12 mutant worms showed significantly lower levels of chemotaxis to odorants compared to wild-type worms throughout their lifespan, and this deficit increased with age. The chemotaxis phenotype in sel-12 mutant worms is rescued by transgenic over-expression of human wild-type PS1, but not the classic fAD-associated variant PS1C410Y, when expression was driven by either the endogenous sel-12 promoter (Psel-12), a pan-neuronal promoter (Primb-1), or by a promoter whose primary expression was in the sensory neurons responsible for the chemotaxis behavior (Psra-6, Podr-10). The behavioural phenotype was also rescued by over-expressing an atypical fAD-linked mutation in PS1 (PS1ΔS169) that has been reported to leave the Notch pathway intact. An examination of the morphology of polymodal nociceptive (ASH) neurons responsible for the chemotaxis behavior also showed increased neurodegeneration over time in sel-12 mutant worms that could be rescued by the same transgenes that rescued the behaviour, demonstrating a parallel with the observed behavioral deficits. Thus, we report an Aß-independent neurodegeneration in C. elegans that was rescued by cell specific over-expression of wild-type human presenilin.
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Enfermedad de Alzheimer , Péptidos beta-Amiloides , Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Mutación , Presenilina-1 , Animales , Humanos , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/metabolismo , Animales Modificados Genéticamente , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Quimiotaxis/genética , Modelos Animales de Enfermedad , Presenilina-1/genética , Células Receptoras Sensoriales/metabolismo , Células Receptoras Sensoriales/patologíaRESUMEN
Large vesicle extrusion from neurons may contribute to spreading pathogenic protein aggregates and promoting inflammatory responses, two mechanisms leading to neurodegenerative disease. Factors that regulate the extrusion of large vesicles, such as exophers produced by proteostressed C. elegans touch neurons, are poorly understood. Here, we document that mechanical force can significantly potentiate exopher extrusion from proteostressed neurons. Exopher production from the C. elegans ALMR neuron peaks at adult day 2 or 3, coinciding with the C. elegans reproductive peak. Genetic disruption of C. elegans germline, sperm, oocytes, or egg/early embryo production can strongly suppress exopher extrusion from the ALMR neurons during the peak period. Conversely, restoring egg production at the late reproductive phase through mating with males or inducing egg retention via genetic interventions that block egg-laying can strongly increase ALMR exopher production. Overall, genetic interventions that promote ALMR exopher production are associated with expanded uterus lengths and genetic interventions that suppress ALMR exopher production are associated with shorter uterus lengths. In addition to the impact of fertilized eggs, ALMR exopher production can be enhanced by filling the uterus with oocytes, dead eggs, or even fluid, supporting that distention consequences, rather than the presence of fertilized eggs, constitute the exopher-inducing stimulus. We conclude that the mechanical force of uterine occupation potentiates exopher extrusion from proximal proteostressed maternal neurons. Our observations draw attention to the potential importance of mechanical signaling in extracellular vesicle production and in aggregate spreading mechanisms, making a case for enhanced attention to mechanobiology in neurodegenerative disease.
Neurons are specialized cells in the brain and nervous system that transmit signals between the brain and the rest of the body, enabling humans and animals to react to internal and external stimuli. For this communication system to function effectively, neurons must remain healthy. Neurons maintain their function in a variety of ways, including by removing excess or damaged cellular components (such as organelles and protein aggregates) that could compromise neuron function. One way to do this is by extruding organelles and aggregates. During 'extrusion events', the material to be removed is gathered within a budding portion of the plasma membrane, which forms a vesicle that ejects the material from the neuron. However, the factors driving the extrusion process remained unknown. To investigate, Wang, Guasp, Salam et al. conducted experiments in the roundworm Caenorhabditis elegans, finding that the number of extrusion events in a certain type of neuron increases at the peak of reproduction. More specifically, a greater number of extrusion events were associated with the presence of fertilized eggs, which accumulate in the uterus before they are laid. Disrupting eggs, sperm or the fertilization process suppressed the increase in extrusion events, suggesting the presence of fertilized eggs is responsible. To determine how the eggs might trigger extrusion events, Wang et al. stretched the uterus using dead eggs, unfertilized eggs or by injecting fluid, finding that each of these approaches increased the number of extrusion events. Further analysis suggests that this mechanical stretching of the uterus signals to the neurons that reproduction has started, encouraging the neurons to remove old components and optimize their function. Wang et al. hypothesize that this stretch response could support neuronal behaviors that aid in successful reproduction, such as sensing food and selecting where to lay eggs. The findings increase our understanding of the factors that trigger vesicle extrusion in living organisms. These observations could have implications for human neurodegenerative diseases such as Alzheimer's disease, in which protein aggregates accumulate in neurons. It is possible that mechanical signals generated by factors associated with Alzheimer's disease, such as high blood pressure, could influence neuronal extrusion and contribute to some of the mechanisms underlying aggregate transfer in neurodegenerative diseases.
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Caenorhabditis elegans , Neuronas , Animales , Caenorhabditis elegans/fisiología , Neuronas/metabolismo , Neuronas/fisiología , Femenino , Útero/metabolismo , Útero/fisiología , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , MasculinoRESUMEN
Chemical mutagenesis-driven forward genetic screens are pivotal in unveiling gene functions, yet identifying causal mutations behind phenotypes remains laborious, hindering their high-throughput application. Here, we reveal a non-uniform mutation rate caused by Ethyl Methane Sulfonate (EMS) mutagenesis in the C. elegans genome, indicating that mutation frequency is influenced by proximate sequence context and chromatin status. Leveraging these factors, we developed a machine learning enhanced pipeline to create a comprehensive EMS mutagenesis probability map for the C. elegans genome. This map operates on the principle that causative mutations are enriched in genetic screens targeting specific phenotypes among random mutations. Applying this map to Whole Genome Sequencing (WGS) data of genetic suppressors that rescue a C. elegans ciliary kinesin mutant, we successfully pinpointed causal mutations without generating recombinant inbred lines. This method can be adapted in other species, offering a scalable approach for identifying causal genes and revitalizing the effectiveness of forward genetic screens.
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Caenorhabditis elegans , Metanosulfonato de Etilo , Aprendizaje Automático , Mutagénesis , Mutación , Caenorhabditis elegans/genética , Animales , Fenotipo , Secuenciación Completa del Genoma/métodos , Cinesinas/genética , Tasa de Mutación , Proteínas de Caenorhabditis elegans/genética , Mapeo Cromosómico/métodosRESUMEN
Background: Recent advances linking gut dysbiosis with neurocognitive disorders such as Alzheimer's disease (AD) suggest that the microbiota-gut-brain axis could be targeted for AD prevention, management, or treatment. Objective: We sought to identify probiotics that can delay Aß-induced paralysis. Methods: Using C. elegans expressing human amyloid-ß (Aß)1-42 in body wall muscles (GMC101), we assessed the effects of several probiotic strains on paralysis. Results: We found that Lacticaseibacillus rhamnosus HA-114 and Bacillus subtilis R0179, but not their supernatants or heat-treated forms, delayed paralysis and prolonged lifespan without affecting the levels of amyloid-ß aggregates. To uncover the mechanism involved, we explored the role of two known pathways involved in neurogenerative diseases, namely mitophagy, via deletion of the mitophagy factor PINK-1, and fatty acid desaturation, via deletion of the Δ9 desaturase FAT-5. Pink-1 deletion in GMC101 worms did not modify the life-prolonging and anti-paralysis effects of HA-114 but reduced the protective effect of R0179 against paralysis without affecting its life-prolonging effect. Upon fat5 deletion in GMC101 worms, the monounsaturated C14:1 and C16:1 FAs conserved their beneficial effect while the saturated C14:0 and C16:0 FAs did not. The beneficial effects of R0179 on both lifespan and paralysis remained unaffected by fat-5 deletion, while the beneficial effect of HA-114 on paralysis and lifespan was significantly reduced. Conclusions: Collectively with clinical and preclinical evidence in other models, our results suggest that HA-114 or R0179 could be studied as potential therapeutical adjuncts in neurodegenerative diseases such as AD.
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Péptidos beta-Amiloides , Bacillus subtilis , Caenorhabditis elegans , Lacticaseibacillus rhamnosus , Longevidad , Probióticos , Animales , Longevidad/efectos de los fármacos , Péptidos beta-Amiloides/metabolismo , Péptidos beta-Amiloides/toxicidad , Parálisis , Fragmentos de Péptidos/toxicidad , Fragmentos de Péptidos/metabolismo , Animales Modificados Genéticamente , Humanos , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismoRESUMEN
The preservation of genome integrity during sperm and egg development is vital for reproductive success. During meiosis, the tumor suppressor BRCA1/BRC-1 and structural maintenance of chromosomes 5/6 (SMC-5/6) complex genetically interact to promote high fidelity DNA double strand break (DSB) repair, but the specific DSB repair outcomes these proteins regulate remain unknown. Using genetic and cytological methods to monitor resolution of DSBs with different repair partners in Caenorhabditis elegans, we demonstrate that both BRC-1 and SMC-5 repress intersister crossover recombination events. Sequencing analysis of conversion tracts from homolog-independent DSB repair events further indicates that BRC-1 regulates intersister/intrachromatid noncrossover conversion tract length. Moreover, we find that BRC-1 specifically inhibits error prone repair of DSBs induced at mid-pachytene. Finally, we reveal functional interactions of BRC-1 and SMC-5/6 in regulating repair pathway engagement: BRC-1 is required for localization of recombinase proteins to DSBs in smc-5 mutants and enhances DSB repair defects in smc-5 mutants by repressing theta-mediated end joining (TMEJ). These results are consistent with a model in which some functions of BRC-1 act upstream of SMC-5/6 to promote recombination and inhibit error-prone DSB repair, while SMC-5/6 acts downstream of BRC-1 to regulate the formation or resolution of recombination intermediates. Taken together, our study illuminates the coordinated interplay of BRC-1 and SMC-5/6 to regulate DSB repair outcomes in the germline.
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Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Roturas del ADN de Doble Cadena , Reparación del ADN , Meiosis , Animales , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Intercambio Genético , Proteína BRCA1/metabolismo , Proteína BRCA1/genéticaRESUMEN
The MUT-7 family of 3'-5' exoribonucleases is evolutionarily conserved across the animal kingdom and plays essential roles in small RNA production in the germline. Most MUT-7 homologues carry a C-terminal domain of unknown function named MUT7-C appended to the exoribonuclease domain. Our analysis shows that the MUT7-C is evolutionary ancient, as a minimal version of the domain exists as an individual protein in prokaryotes. In animals, MUT7-C has acquired an insertion that diverged during evolution, expanding its functions. Caenorhabditis elegans MUT-7 contains a specific insertion within MUT7-C, which allows binding to MUT-8 and, consequently, MUT-7 recruitment to germ granules. In addition, in C. elegans and human MUT-7, the MUT7-C domain contributes to RNA binding and is thereby crucial for ribonuclease activity. This RNA-binding function most likely represents the ancestral function of the MUT7-C domain. Overall, this study sheds light on MUT7-C and assigns two functions to this previously uncharacterized domain.
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Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Exorribonucleasas , Dominios Proteicos , Animales , Exorribonucleasas/metabolismo , Exorribonucleasas/química , Exorribonucleasas/genética , Caenorhabditis elegans/enzimología , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/química , Humanos , Evolución Molecular , ARN/metabolismo , ARN/química , Secuencia de Aminoácidos , Unión ProteicaRESUMEN
Parkinson's disease (PD) is a common neurodegenerative disorder characterized by symptoms such as bradykinesia, resting tremor, and rigidity, primarily driven by the degradation of dopaminergic (DA) neurons in the substantia nigra. A significant contributor to familial autosomal dominant PD cases is mutations in the LRRK2 gene, making it a primary therapeutic target. This study explores the role of microRNAs (miRNAs) in regulating the proteomic stress responses associated with neurodegeneration in PD using C. elegans models. Our focus is on miR-71, a miRNA known to affect stress resistance and act as a pro-longevity factor in C. elegans. We investigated miR-71's function in C. elegans models of PD, where mutant LRRK2 expression correlates with dopaminergic neuronal death. Our findings reveal that miR-71 overexpression rescues motility defects and slows dopaminergic neurodegeneration in these models, suggesting its critical role in mitigating the proteotoxic effects of mutant LRRK2. Conversely, miR-71 knockout exacerbates neuronal death caused by mutant LRRK2. Additionally, our data indicate that miR-71's neuroprotective effect involves downregulating the toll receptor domain protein tir-1, implicating miR-71 repression of tir-1 as vital in the response to LRRK2-induced proteotoxicity. These insights into miR-71's role in C. elegans models of PD not only enhance our understanding of molecular mechanisms in neurodegeneration but also pave the way for potential research into human neurodegenerative diseases, leveraging the conservation of miRNAs and their targets across species.