RESUMEN
Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder, of the so-called minority diseases, due to its low prevalence. It is caused by an abnormally long track of glutamines (polyQs) in mutant huntingtin (mHtt), which makes the protein toxic and prone to aggregation. Many pathways of clearance of badly-folded proteins are disrupted in neurons of patients with HD. In this work, we show that one Mn(II) quinone complex (4QMn), designed to work as an artificial superoxide dismutase, is able to activate both the ubiquitin-proteasome system and the autophagy pathway in vitro and in vivo models of HD. Activation of these pathways degrades mHtt and other protein-containing polyQs, which restores proteostasis in these models. Hence, we propose 4QMn as a potential drug to develop a therapy to treat HD.
Asunto(s)
Enfermedad de Huntington , Quinolinas , Animales , Modelos Animales de Enfermedad , Humanos , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Enfermedad de Huntington/tratamiento farmacológico , Enfermedad de Huntington/metabolismo , Manganeso , Modelos Teóricos , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteostasis , Quinolinas/uso terapéuticoRESUMEN
RNA interference (RNAi) is a widespread and widely exploited phenomenon. Here, we show that changing inositol 1,4,5-trisphosphate (IP3) signalling alters RNAi sensitivity in Caenorhabditis elegans. Reducing IP3 signalling enhances sensitivity to RNAi in a broad range of genes and tissues. Conversely up-regulating IP3 signalling decreases sensitivity. Tissue-specific rescue experiments suggest IP3 functions in the intestine. We also exploit IP3 signalling mutants to further enhance the sensitivity of RNAi hypersensitive strains. These results demonstrate that conserved cell signalling pathways can modify RNAi responses, implying that RNAi responses may be influenced by an animal's physiology or environment.
Asunto(s)
Caenorhabditis elegans/fisiología , Inositol 1,4,5-Trifosfato/metabolismo , Interferencia de ARN/fisiología , Transducción de Señal/fisiología , Animales , Caenorhabditis elegans/genética , Procesamiento de Imagen Asistido por Computador , Mucosa Intestinal/metabolismo , Microscopía Fluorescente , Modelos Biológicos , ARN Bicatenario , Transducción de Señal/genéticaRESUMEN
Huntington disease is a neurodegenerative condition for which there is no cure to date. Activation of AMP-activated protein kinase has previously been shown to be beneficial in in vitro and in vivo models of Huntington's disease. Moreover, a recent cross-sectional study demonstrated that treatment with metformin, a well-known activator of this enzyme, is associated with better cognitive scores in patients with this disease. We performed a preclinical study using metformin to treat phenotypes of the zQ175 mouse model of Huntington disease. We evaluated behavior (motor and neuropsychiatric function) and molecular phenotypes (aggregation of mutant huntingtin, levels of brain-derived neurotrophic factor, neuronal inflammation, etc.). We also used two models of polyglutamine toxicity in Caenorhabditis elegans to further explore potential mechanisms of metformin action. Our results provide strong evidence that metformin alleviates motor and neuropsychiatric phenotypes in zQ175 mice. Moreover, metformin intake reduces the number of nuclear aggregates of mutant huntingtin in the striatum. The expression of brain-derived neurotrophic factor, which is reduced in mutant animals, is partially restored in metformin-treated mice, and glial activation in mutant mice is reduced in metformin-treated animals. In addition, using worm models of polyglutamine toxicity, we demonstrate that metformin reduces polyglutamine aggregates and restores neuronal function through mechanisms involving AMP-activated protein kinase and lysosomal function. Our data indicate that metformin alleviates the progression of the disease and further supports AMP-activated protein kinase as a druggable target against Huntington's disease.
Asunto(s)
Enfermedad de Huntington/tratamiento farmacológico , Hipoglucemiantes/uso terapéutico , Metformina/uso terapéutico , Agregación Patológica de Proteínas/tratamiento farmacológico , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patología , Caenorhabditis elegans , Modelos Animales de Enfermedad , Humanos , Proteína Huntingtina/metabolismo , Enfermedad de Huntington/metabolismo , Enfermedad de Huntington/patología , Ratones , Péptidos/metabolismo , Agregación Patológica de Proteínas/metabolismo , Agregación Patológica de Proteínas/patologíaRESUMEN
In the presence of aggregation-prone proteins, the cytosol and endoplasmic reticulum (ER) undergo a dramatic shift in their respective redox status, with the cytosol becoming more oxidized and the ER more reducing. However, whether and how changes in the cellular redox status may affect protein aggregation is unknown. Here, we show that C. elegans loss-of-function mutants for the glutathione reductase gsr-1 gene enhance the deleterious phenotypes of heterologous human, as well as endogenous worm aggregation-prone proteins. These effects are phenocopied by the GSH-depleting agent diethyl maleate. Additionally, gsr-1 mutants abolish the nuclear translocation of HLH-30/TFEB transcription factor, a key inducer of autophagy, and strongly impair the degradation of the autophagy substrate p62/SQST-1::GFP, revealing glutathione reductase may have a role in the clearance of protein aggregates by autophagy. Blocking autophagy in gsr-1 worms expressing aggregation-prone proteins results in strong synthetic developmental phenotypes and lethality, supporting the physiological importance of glutathione reductase in the regulation of misfolded protein clearance. Furthermore, impairing redox homeostasis in both yeast and mammalian cells induces toxicity phenotypes associated with protein aggregation. Together, our data reveal that glutathione redox homeostasis may be central to proteostasis maintenance through autophagy regulation.
Asunto(s)
Autofagia/genética , Caenorhabditis elegans/genética , Glutatión Reductasa/metabolismo , Glutatión/metabolismo , Péptidos/toxicidad , Agregación Patológica de Proteínas/metabolismo , Proteostasis/genética , Péptidos beta-Amiloides/genética , Péptidos beta-Amiloides/metabolismo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Caenorhabditis elegans/crecimiento & desarrollo , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Línea Celular , Retículo Endoplásmico/metabolismo , Glutatión/genética , Glutatión Reductasa/genética , Homeostasis/efectos de los fármacos , Homeostasis/genética , Humanos , Maleatos/farmacología , Células Musculares/metabolismo , Neuronas/metabolismo , Oxidación-Reducción/efectos de los fármacos , Péptidos/antagonistas & inhibidores , Fenotipo , Proteolisis/efectos de los fármacos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteína Sequestosoma-1/genética , Proteína Sequestosoma-1/metabolismo , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismoRESUMEN
Huntington's disease (HD) is an inherited, dominant neurodegenerative disorder caused by an abnormal expansion of CAG triplets in the huntingtin gene (htt). Despite extensive efforts to modify the progression of HD thus far only symptomatic treatment is available. Recent work suggests that treating invertebrate and mice HD models with metformin, a well-known AMPK activator which is used worldwide to treat type 2-diabetes, reduces mutant huntingtin from cells and alleviates many of the phenotypes associated to HD. Herein we report statistical analyses of a sample population of participants in the Enroll-HD database, a world-wide observational study on HD, to assess the effect of metformin intake in HD patients respect to cognitive status using linear models. This cross-sectional study shows for the first time that the use of metformin associates with better cognitive function in HD patients.
Asunto(s)
Cognición/fisiología , Enfermedad de Huntington/tratamiento farmacológico , Hipoglucemiantes/uso terapéutico , Metformina/uso terapéutico , Adulto , Anciano , Estudios de Casos y Controles , Cognición/efectos de los fármacos , Estudios Transversales , Femenino , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Usher syndrome type I (USH1) is an autosomal recessive disorder characterized by congenital profound deafness, vestibular areflexia and prepubertal retinitis pigmentosa. The first purpose of this study was to determine the pathologic nature of eighteen USH1 putative splicing variants found in our series and their effect in the splicing process by minigene assays. These variants were selected according to bioinformatic analysis. The second aim was to analyze the USH1 transcripts, obtained from nasal epithelial cells samples of our patients, in order to corroborate the observed effect of mutations by minigenes in patient's tissues. The last objective was to evaluate the nasal ciliary beat frequency in patients with USH1 and compare it with control subjects. In silico analysis were performed using four bioinformatic programs: NNSplice, Human Splicing Finder, NetGene2 and Spliceview. Afterward, minigenes based on the pSPL3 vector were used to investigate the implication of selected changes in the mRNA processing. To observe the effect of mutations in the patient's tissues, RNA was extracted from nasal epithelial cells and RT-PCR analyses were performed. Four MYO7A (c.470G>A, c.1342_1343delAG, c.5856G>A and c.3652G>A), three CDH23 (c.2289+1G>A, c.6049G>A and c.8722+1delG) and one PCDH15 (c.3717+2dupTT) variants were observed to affect the splicing process by minigene assays and/or transcripts analysis obtained from nasal cells. Based on our results, minigenes are a good approach to determine the implication of identified variants in the mRNA processing, and the analysis of RNA obtained from nasal epithelial cells is an alternative method to discriminate neutral Usher variants from those with a pathogenic effect on the splicing process. In addition, we could observe that the nasal ciliated epithelium of USH1 patients shows a lower ciliary beat frequency than control subjects.