RESUMEN
DJ-1 is an oxidation sensitive protein encoded by the PARK7 gene. Mutations in PARK7 are a rare cause of familial recessive Parkinson's disease (PD), but growing evidence suggests involvement of DJ-1 in idiopathic PD. The key clinical features of PD, rigidity and bradykinesia, result from neurotransmitter imbalance, particularly the catecholamines dopamine (DA) and noradrenaline. We report in human brain and human SH-SY5Y neuroblastoma cell lines that DJ-1 predominantly forms high molecular weight (HMW) complexes that included RNA metabolism proteins hnRNPA1 and PABP1 and the glycolysis enzyme GAPDH. In cell culture models the oxidation status of DJ-1 determined the specific complex composition. RNA sequencing indicated that oxidative changes to DJ-1 were concomitant with changes in mRNA transcripts mainly involved in catecholamine metabolism. Importantly, loss of DJ-1 function upon knock down (KD) or expression of the PD associated form L166P resulted in the absence of HMW DJ-1 complexes. In the KD model, the absence of DJ-1 complexes was accompanied by impairment in catecholamine homeostasis, with significant increases in intracellular DA and noraderenaline levels. These changes in catecholamines could be rescued by re-expression of DJ-1. This catecholamine imbalance may contribute to the particular vulnerability of dopaminergic and noradrenergic neurons to neurodegeneration in PARK7-related PD. Notably, oxidised DJ-1 was significantly decreased in idiopathic PD brain, suggesting altered complex function may also play a role in the more common sporadic form of the disease.
Asunto(s)
Catecolaminas/metabolismo , Proteína Desglicasa DJ-1/genética , Proteína Desglicasa DJ-1/metabolismo , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Encéfalo/metabolismo , Línea Celular Tumoral , Dopamina/metabolismo , Homeostasis , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Oxidación-Reducción , Estrés Oxidativo/fisiología , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/metabolismoRESUMEN
Mutations in polycystin-1 and transient receptor potential polycystin 2 (TRPP2) account for almost all clinically identified cases of autosomal dominant polycystic kidney disease (ADPKD), one of the most common human genetic diseases. TRPP2 functions as a cation channel in its homomeric complex and in the TRPP2/polycystin-1 receptor/ion channel complex. The activation mechanism of TRPP2 is unknown, which significantly limits the study of its function and regulation. Here, we generated a constitutively active gain-of-function (GOF) mutant of TRPP2 by applying a mutagenesis scan on the S4-S5 linker and the S5 transmembrane domain, and studied functional properties of the GOF TRPP2 channel. We found that extracellular divalent ions, including Ca(2+), inhibit the permeation of monovalent ions by directly blocking the TRPP2 channel pore. We also found that D643, a negatively charged amino acid in the pore, is crucial for channel permeability. By introducing single-point ADPKD pathogenic mutations into the GOF TRPP2, we showed that different mutations could have completely different effects on channel activity. The in vivo function of the GOF TRPP2 was investigated in zebrafish embryos. The results indicate that, compared with wild type (WT), GOF TRPP2 more efficiently rescued morphological abnormalities, including curly tail and cyst formation in the pronephric kidney, caused by down-regulation of endogenous TRPP2 expression. Thus, we established a GOF TRPP2 channel that can serve as a powerful tool for studying the function and regulation of TRPP2. The GOF channel may also have potential application for developing new therapeutic strategies for ADPKD.
Asunto(s)
Canales Catiónicos TRPP/fisiología , Amilorida/farmacología , Animales , Calcio/metabolismo , Técnicas de Silenciamiento del Gen , Humanos , Mutación/genética , Mutación Puntual/genética , Riñón Poliquístico Autosómico Dominante/genética , Rojo de Rutenio/farmacología , Canales Catiónicos TRPP/efectos de los fármacos , Canales Catiónicos TRPP/genética , Pez Cebra/embriologíaRESUMEN
The last decade has experienced the emergence of microRNAs as a key molecular tool for the diagnosis and prognosis of human diseases. Although the focus has mostly been on cancer, neurodegenerative diseases present an exciting, yet less explored, platform for microRNA research. Several studies have highlighted the significance of microRNAs in neurogenesis and neurodegeneration, and pre-clinical studies have shown the potential of microRNAs as biomarkers. Despite this, no bona fide microRNAs have been identified as true diagnostic or prognostic biomarkers for neurodegenerative disease. This is mainly due to the lack of precisely defined patient cohorts and the variability within and between individual cohorts. However, the discovery that microRNAs exist as stable molecules at detectable levels in body fluids has opened up new avenues for microRNAs as potential biomarker candidates. Furthermore, technological developments in microRNA biology have contributed to the possible design of microRNA-mediated disease intervention strategies. The combination of these advancements, with the availability of well-defined longitudinal patient cohort, promises to not only assist in developing invaluable diagnostic tools for clinicians, but also to increase our overall understanding of the underlying heterogeneity of neurodegenerative diseases. In this review, we present a comprehensive overview of the existing knowledge of microRNAs in neurodegeneration and provide a perspective of the applicability of microRNAs as a basis for future therapeutic intervention strategies.
Asunto(s)
MicroARNs/genética , MicroARNs/uso terapéutico , Enfermedades Neurodegenerativas/genética , Enfermedades Neurodegenerativas/terapia , Animales , Autofagia , Encéfalo/metabolismo , Encéfalo/patología , Descubrimiento de Drogas , Regulación de la Expresión Génica , Marcadores Genéticos , Terapia Genética , Humanos , MicroARNs/análisis , MicroARNs/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Enfermedades Neurodegenerativas/patología , Transcripción GenéticaRESUMEN
Although mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most common cause of genetic Parkinson's disease, their function is largely unknown. LRRK2 is pleiotropic in nature, shown to be involved in neurodegeneration and in more peripheral processes, including kidney functions, in rats and mice. Recent studies in zebrafish have shown conflicting evidence that removal of the LRRK2 WD40 domain may or may not affect dopaminergic neurons and/or locomotion. This study shows that â¼50% LRRK2 knockdown in zebrafish causes not only neuronal loss but also developmental perturbations such as axis curvature defects, ocular abnormalities, and edema in the eyes, lens, and otic vesicles. We further show that LRRK2 knockdown results in significant neuronal loss, including a reduction of dopaminergic neurons. Immunofluorescence demonstrates that endogenous LRRK2 is expressed in the lens, brain, heart, spinal cord, and kidney (pronephros), which mirror the LRRK2 morphant phenotypes observed. LRRK2 knockdown results further in the concomitant upregulation of ß-synuclein, PARK13, and SOD1 and causes ß-synuclein aggregation in the diencephalon, midbrain, hindbrain, and postoptic commissure. LRRK2 knockdown causes mislocalization of the Na(+) /K(+) ATPase protein in the pronephric ducts, suggesting that the edema might be linked to renal malfunction and that LRRK2 might be associated with pronephric duct epithelial cell differentiation. Combined, our study shows that LRRK2 has multifaceted roles in zebrafish and that zebrafish represent a complementary model to further our understanding of this central protein. © 2016 Wiley Periodicals, Inc.
Asunto(s)
Anomalías Múltiples/genética , Anomalías Múltiples/patología , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina/genética , Enfermedades Neurodegenerativas/genética , Neuronas/patología , Proteínas de Pez Cebra/genética , Sinucleína beta/genética , Secuencia de Aminoácidos , Animales , Química Encefálica/genética , Neuronas Dopaminérgicas , Técnicas de Silenciamiento del Gen , Humanos , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina/biosíntesis , Locomoción , Mutación/genética , Enfermedades Neurodegenerativas/patología , Enfermedad de Parkinson/genética , ATPasa Intercambiadora de Sodio-Potasio/genética , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Pez Cebra , Proteínas de Pez Cebra/biosíntesisRESUMEN
The widespread use of herbicides and antibiotics for selection of transgenic plants has not been very successful with regard to commercialization and public acceptance. Hence, alternative selection systems are required. In this study, we describe the use of ipt, the bacterial gene encoding the enzyme isopentenyl transferase from Agrobacterium tumefaciens, as a positive selectable marker for plastid transformation. A comparison between the traditional spectinomycin-based aadA selection system and the ipt selection system demonstrated that selection of transplastomic plants on medium lacking cytokinin was as effective as selection on medium containing spectinomycin. Proof of principle was demonstrated by transformation of the kasIII gene encoding 3-ketoacyl acyl carrier protein synthase III into tobacco plastids. Transplastomic tobacco plants were readily obtained using the ipt selection system, and were phenotypically normal despite over-expression of isopentenyl transferase. Over-expression of KASIII resulted in a significant increase in 16:0 fatty acid levels, and a significant decrease in the levels of 18:0 and 18:1 fatty acids. Our study demonstrates use of a novel positive plastid transformation system that may be used for selection of transplastomic plants without affecting the expression of transgenes within the integrated vector cassette or the resulting activity of the encoded protein. This system has the potential to be applied to monocots, which are typically not amenable to traditional antibiotic-based selection systems, and may be used in combination with a negative selectable marker as part of a two-step selection system to obtain homoplasmic plant lines.
Asunto(s)
Citocininas/metabolismo , Ácidos Grasos/metabolismo , Nicotiana/metabolismo , Espectinomicina/farmacología , Transferasas Alquil y Aril/genética , Transferasas Alquil y Aril/metabolismo , Expresión Génica , Vectores Genéticos , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente , Plastidios/metabolismo , Nicotiana/genética , Transformación Genética , TransgenesRESUMEN
The progressive loss of motor control due to reduction of dopamine-producing neurons in the substantia nigra pars compacta and decreased striatal dopamine levels are the classically described features of Parkinson disease (PD). Neuronal damage also progresses to other regions of the brain, and additional non-motor dysfunctions are common. Accumulation of environmental toxins, such as pesticides and metals, are suggested risk factors for the development of typical late onset PD, although genetic factors seem to be substantial in early onset cases. Mutations of DJ-1 are known to cause a form of recessive early onset Parkinson disease, highlighting an important functional role for DJ-1 in early disease prevention. This study identifies human DJ-1 as a metal-binding protein able to evidently bind copper as well as toxic mercury ions in vitro. The study further characterizes the cytoprotective function of DJ-1 and PD-mutated variants of DJ-1 with respect to induced metal cytotoxicity. The results show that expression of DJ-1 enhances the cells' protective mechanisms against induced metal toxicity and that this protection is lost for DJ-1 PD mutations A104T and D149A. The study also shows that oxidation site-mutated DJ-1 C106A retains its ability to protect cells. We also show that concomitant addition of dopamine exposure sensitizes cells to metal-induced cytotoxicity. We also confirm that redox-active dopamine adducts enhance metal-catalyzed oxidation of intracellular proteins in vivo by use of live cell imaging of redox-sensitive S3roGFP. The study indicates that even a small genetic alteration can sensitize cells to metal-induced cell death, a finding that may revive the interest in exogenous factors in the etiology of PD.
Asunto(s)
Cobre/toxicidad , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Mercurio/toxicidad , Proteínas Oncogénicas/metabolismo , Enfermedad de Parkinson/metabolismo , Animales , Secuencia de Bases , Línea Celular , Cartilla de ADN , Dopamina/farmacología , Homeostasis , Humanos , Péptidos y Proteínas de Señalización Intracelular/química , Péptidos y Proteínas de Señalización Intracelular/genética , Ratones , Modelos Moleculares , Proteínas Oncogénicas/química , Proteínas Oncogénicas/genética , Oxidación-Reducción , Unión Proteica , Proteína Desglicasa DJ-1RESUMEN
Parkinson's disease (PD) is a progressive and irreversible neurodegenerative disorder coupled to selective degeneration of dopamine-producing neurons in the substantia nigra. The majority of PD incidents are sporadic, but monogenic cases account for 5-10% of cases. Mutations in PINK1 cause autosomal recessive forms of early-onset PD, and PINK1 stimulates Omi/HtrA2/PARK13 protease activity when both proteins act as neuroprotective components in the same stress pathway. Studies on PINK1 and PARK13 have concentrated on phosphorylation-dependent PINK1-mediated activation of PARK13 and mitochondrial functions, because both proteins are classically viewed as mitochondrial. Although PARK13-mediated protective mechanisms are at least in part regulated by PINK1, little is known concerning how these two proteins are regulated in different subcellular compartments or, indeed, the influence of PARK13 on PINK1 characteristics. We show that PARK13 localizes to a variety of subcellular locations in neuronal cells and that PINK1, although more restrictive, also localizes to locations other than those previously reported. We demonstrate that PARK13 accumulation leads to a concomitant accumulation of PINK1 and that the increase in PINK1 levels is compartmental specific, indicating a correlative relationship between the two proteins. Moreover, we show that PARK13 and PINK1 protein levels accumulate in response to H2 O2 and L-DOPA treatments in a subcellular fashion and that both proteins show relocation to the cytoskeleton in response to H2 O2 . This H2 O2 -mediated relocation is abolished by PARK13 overexpression. This study shows that PARK13 and PINK1 are subcellular-specific, but dynamic, proteins with a reciprocal molecular relationship providing new insight into the complexity of PD.
Asunto(s)
Neuronas/citología , Estrés Oxidativo/fisiología , Enfermedad de Parkinson/metabolismo , Proteínas Quinasas/metabolismo , Fracciones Subcelulares/metabolismo , Línea Celular Tumoral , Citoesqueleto/efectos de los fármacos , Citoesqueleto/metabolismo , Dopaminérgicos/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/genética , Humanos , Peróxido de Hidrógeno/farmacología , Levodopa/farmacología , Mutación/genética , Neuroblastoma/patología , Neuronas/efectos de los fármacos , Oxidantes/farmacología , Estrés Oxidativo/efectos de los fármacos , Enfermedad de Parkinson/genética , Proteínas Quinasas/genética , ARN Mensajero/metabolismo , Fracciones Subcelulares/efectos de los fármacos , Factores de Tiempo , TransfecciónRESUMEN
Understanding the complex biology of the brain requires analyzing its structural and functional complexity at the protein level. The large-scale analysis of the brain proteome, coupled with characterization of central brain proteins, provides insight into fundamental brain processes and processes linked to neurodegenerative diseases. Here we provide a map of the zebrafish brain proteome by using two-dimensional gel electrophoresis (2DE), followed by the identification of 95 brain proteins using mass spectrometry (LC-ESI MS/MS). Our data show extensive phosphorylation of brain proteins but less prominent glycosylation. Furthermore, ~51% of the identified proteins are predicted to have one or more ubiquitination sites whereas ~90% are predicted to have one or more SUMOylation sites. Our findings provide a valuable proteome map of the zebrafish brain and associated posttranslational modifications demonstrating that zebrafish proteomic approaches can aid in our understanding of proteins central to important neuronal processes and those associated with neurodegenerative disorders.
Asunto(s)
Encéfalo/metabolismo , Degeneración Nerviosa/metabolismo , Proteoma/metabolismo , Proteínas de Pez Cebra/metabolismo , Animales , Electroforesis en Gel Bidimensional , Glicosilación , Degeneración Nerviosa/genética , Fosforilación , Proteoma/genética , Proteómica/métodos , Espectrometría de Masas en Tándem , Pez Cebra/genética , Pez Cebra/metabolismo , Proteínas de Pez Cebra/genéticaRESUMEN
BACKGROUND: Parkinson's disease (PD) is the second most common neurodegenerative movement disorder, caused by preferential dopaminergic neuronal cell death in the substantia nigra, a process also influenced by oxidative stress. L-3,4-dihydroxyphenylalanine (L-DOPA) represents the main treatment route for motor symptoms associated with PD however, its exact mode of action remains unclear. A spectrum of conflicting data suggests that L-DOPA may damage dopaminergic neurons due to oxidative stress whilst other data suggest that L-DOPA itself may induce low levels of oxidative stress, which in turn stimulates endogenous antioxidant mechanisms and neuroprotection. RESULTS: In this study we performed a two-dimensional gel electrophoresis (2DE)-based proteomic study to gain further insight into the mechanism by which L-DOPA can influence the toxic effects of H2O2 in neuronal cells. We observed that oxidative stress affects metabolic pathways as well as cytoskeletal integrity and that neuronal cells respond to oxidative conditions by enhancing numerous survival pathways. Our study underlines the complex nature of L-DOPA in PD and sheds light on the interplay between oxidative stress and L-DOPA. CONCLUSIONS: Oxidative stress changes neuronal metabolic routes and affects cytoskeletal integrity. Further, L-DOPA appears to reverse some H2O2-mediated effects evident at both the proteome and cellular level.
Asunto(s)
Antiparkinsonianos/farmacología , Levodopa/farmacología , Neuronas/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Proteoma/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Citoesqueleto/efectos de los fármacos , Citoesqueleto/fisiología , Electroforesis en Gel Bidimensional , Humanos , Peróxido de Hidrógeno/toxicidad , Espectrometría de Masas , Neuronas/patología , Neuronas/fisiología , Estrés Oxidativo/fisiología , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Chloroplasts are descendants of cyanobacteria and divide by binary fission. The number of chloroplasts is regulated in a cell type-specific manner to ensure that specialized cell types can perform their functions optimally. Several protein components of the chloroplast division apparatus have been identified in the past several years, but how this process is regulated in response to developmental status, environmental signals and stress is still unknown. To begin to address this we undertook a proteomic analysis of three accumulation and replication of chloroplasts mutants that show a spectrum of plastid division perturbations. We show that defects in the chloroplast division process results in changes in the abundance of proteins when compared to wild type, but that the profile of the native stromal and membrane complexes remains unchanged. Furthermore, by combining BN-PAGE with protein interaction assays we show that AtFtsZ2-1 and AtFtsZ2-2 assemble together with rpl12A and EF-Tu into a novel chloroplast membrane complex.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Cloroplastos/metabolismo , Cloroplastos/metabolismo , Proteoma , Secuencia de Aminoácidos , Arabidopsis/genética , Arabidopsis/fisiología , Cloroplastos/fisiología , Cromatografía Líquida de Alta Presión , Regulación de la Expresión Génica de las Plantas , Mutación , Factor Tu de Elongación Peptídica/metabolismo , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Hojas de la Planta/fisiología , Plantas Modificadas Genéticamente , Proteómica , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Plantones/genética , Plantones/metabolismo , Plantones/fisiología , Espectrometría de Masas en Tándem , Nicotiana/citología , Nicotiana/genética , Nicotiana/metabolismo , Electroforesis Bidimensional Diferencial en Gel , Técnicas del Sistema de Dos HíbridosRESUMEN
Chloroplast division represents a fundamental but complex biological process involving remnants of the ancestral bacterial division machinery and proteins of eukaryotic origin. Moreover, the chloroplast division machinery is divided into stromal and cytosolic sub machineries, which coordinate and control their activities to ensure appropriate division initiation and progression. Dynamin related protein 5B (DRP5B) and plastid division protein 1 and 2 (PDV1 and PDV2) are all plant-derived proteins and represent components of the cytosolic division machinery, where DRP5B is thought to exert constrictional force during division. However, the direct relationship between PDV1, PDV2 and DRP5B, and moreover how DRP5B is regulated during plastid constriction remains unclear. In this study we show that PDV1 and PDV2 can interact with themselves and with each other through their cytosolic domains. We demonstrate that DRP5B interacts with itself and with the cytosolic region of PDV1 and that the two functional isoforms of DRP5B have highly overlapping functions. We further show that DRP5B harbors GTPase activity and moreover that PDV1 and PDV2 inhibits DRP5B-mediated GTP hydrolysis in a ratio dependent manner. Our data suggest that the PDV proteins contribute to the regulation of DRP5B activity thereby enforcing control over the division process during early constriction.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Cloroplastos/genética , Dinaminas/genética , Regulación de la Expresión Génica de las Plantas , Secuencia de Aminoácidos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Sitios de Unión/genética , Cloroplastos/metabolismo , Citosol/metabolismo , Dinaminas/metabolismo , Electroforesis en Gel de Poliacrilamida , GTP Fosfohidrolasas/genética , GTP Fosfohidrolasas/metabolismo , Perfilación de la Expresión Génica , Guanosina Trifosfato/metabolismo , Hidrólisis , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Datos de Secuencia Molecular , Mutación , Plantas Modificadas Genéticamente , Unión Proteica , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Técnicas del Sistema de Dos HíbridosRESUMEN
The Parkinsonism-associated protein DJ-1 has been suggested to activate the Cu-Zn superoxide dismutase (SOD1) by providing its copper cofactor. The structural and chemical means by which DJ-1 could support this function is unknown. In this study, we characterize the molecular interaction of DJ-1 with Cu(I). Mass spectrometric analysis indicates binding of one Cu(I) ion per DJ-1 homodimer. The crystal structure of DJ-1 bound to Cu(I) confirms metal coordination through a docking accessible biscysteinate site formed by juxtaposed cysteine residues at the homodimer interface. Spectroscopy in crystallo validates the identity and oxidation state of the bound metal. The measured subfemtomolar dissociation constant (Kd = 6.41 × 10(-16) M) of DJ-1 for Cu(I) supports the physiological retention of the metal ion. Our results highlight the requirement of a stable homodimer for copper binding by DJ-1. Parkinsonism-linked mutations that weaken homodimer interactions will compromise this capability.
Asunto(s)
Cobre/química , Cisteína/química , Péptidos y Proteínas de Señalización Intracelular/química , Péptidos y Proteínas de Señalización Intracelular/genética , Mutación/fisiología , Proteínas Oncogénicas/química , Proteínas Oncogénicas/genética , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/metabolismo , Sitios de Unión , Cobre/metabolismo , Cristalografía por Rayos X , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Espectrometría de Masas , Metales/química , Modelos Moleculares , Proteínas Oncogénicas/metabolismo , Conformación Proteica , Proteína Desglicasa DJ-1 , Espectrometría de Masa por Ionización de Electrospray , Superóxido Dismutasa/química , Superóxido Dismutasa-1RESUMEN
Plastids are complex organelles that are integrated into the plant host cell where they differentiate and divide in tune with plant differentiation and development. In line with their prokaryotic origin, plastid division involves both evolutionary conserved proteins and proteins of eukaryotic origin where the host has acquired control over the process. The plastid division apparatus is spatially separated between the stromal and the cytosolic space but where clear coordination mechanisms exist between the two machineries. Our knowledge of the plastid division process has increased dramatically during the past decade and recent findings have not only shed light on plastid division enzymology and the formation of plastid division complexes but also on the integration of the division process into a multicellular context. This review summarises our current knowledge of plastid division with an emphasis on biochemical features, the functional assembly of protein complexes and regulatory features of the overall process.
Asunto(s)
Proteínas de Arabidopsis/química , División Celular , Plastidios/química , Arabidopsis/química , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Diferenciación Celular , Proteínas de Cloroplastos/química , Proteínas de Cloroplastos/genética , Citosol/química , Evolución Molecular , Membranas Intracelulares/química , Complejos Multiproteicos/química , Complejos Multiproteicos/genética , Fosforilación , Células Vegetales/química , Plastidios/genéticaRESUMEN
Mutations in the DJ-1 gene (also known as PARK7) cause inherited Parkinson's disease, which is characterized by neuronal death. Although DJ-1 is thought to be an antioxidant protein, the underlying mechanism by which loss of DJ-1 function contributes to cell death is unclear. Human DJ-1 and its Arabidopsis thaliana homologue, AtDJ-1a, are evolutionarily conserved proteins, indicating a universal function. To gain further knowledge of the molecular features associated with DJ-1 dysfunction, we have characterized AtDJ-1a. We show that AtDJ-1a levels are responsive to stress treatment and that AtDJ-1a loss of function results in accelerated cell death in aging plants. By contrast, transgenic plants with elevated AtDJ-1a levels have increased protection against environmental stress conditions, such as strong light, H(2)O(2), methyl viologen and copper sulfate. We further identify superoxide dismutase 1 (SOD1) and glutathione peroxidase 2 (GPX2) as interaction partners of both AtDJ-1a and human DJ-1, and show that this interaction results in AtDJ-1a- and DJ-1-mediated cytosolic SOD1 activation in a copper-dependent fashion. Our data have highlighted a conserved molecular mechanism for DJ-1 and revealed a new protein player in the oxidative stress response of plants.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Superóxido Dismutasa/metabolismo , Apoptosis/genética , Proteínas de Arabidopsis/genética , Clonación Molecular , Secuencia Conservada , Citoprotección , Citosol/metabolismo , Activación Enzimática/genética , Glutatión Peroxidasa/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Luz/efectos adversos , Mutación/genética , Proteínas Oncogénicas/genética , Estrés Oxidativo/genética , Enfermedad de Parkinson/genética , Plantas Modificadas Genéticamente , Unión Proteica , Proteína Desglicasa DJ-1 , Estrés Fisiológico/genética , Superóxido Dismutasa-1 , Transgenes/genéticaRESUMEN
BACKGROUND: : Although the 14-item Starkstein Apathy Scale (SAS) is recommended to screen for and measure the severity of apathetic symptoms in Parkinson disease (PD), the psychometric attributes of this scale have not yet been fully evaluated. OBJECTIVE: : The authors examined the reliability, factor structure, and discriminant validity of the SAS in 194 nondemented patients with early untreated PD. DESIGN: : Cross-sectional multicenter population-based study from Western and Southern Norway. MEASUREMENTS: : Standardized rating scales for parkinsonism and neuropsychiatric symptoms. RESULTS: : The SAS showed fair internal consistency (Cronbach's α = 0.69) and exploratory factor analysis identified two factors: the first factor (24.2% of the variance) represented cognitive-behavioral aspects of apathy (items 1, 2, and 4-8; Cronbach's α = 0.74) and the second factor (15.0% of the variance) a general apathy dimension (items 3 and 9-14; Cronbach's α = 0.52). The correlation between these two factors was low (Spearman's r = 0.19, N = 194, p = 0.008), indicating clinically distinct dimensions, but both factor scores were strongly related to the total SAS score (Spearman's r > 0.6, N = 194, p < 0.0005). Item 3 (insight or self-reflection) showed a negative item-total correlation, and removing this item raised the Cronbach's α of the second factor to 0.70, but did not substantially alter the other results. Both the total score and factor scores of SAS showed fair discriminant validity. CONCLUSIONS: : Although the SAS showed fairly good psychometric properties and the exploratory factor analysis suggested a two-factor solution, the results with this PD sample indicate that item 3 is ambiguous and should be considered removed from the scale.
Asunto(s)
Apatía , Enfermedad de Parkinson/complicaciones , Enfermedad de Parkinson/diagnóstico , Anciano , Estudios Transversales , Análisis Factorial , Femenino , Humanos , Masculino , Persona de Mediana Edad , Psicometría/instrumentación , Reproducibilidad de los Resultados , AutoinformeRESUMEN
Plastids are vital organelles involved in important metabolic functions that directly affect plant growth and development. Plastids divide by binary fission involving the coordination of numerous protein components. A tight control of the plastid division process ensures that: there is a full plastid complement during and after cell division, specialized cell types have optimal plastid numbers; the division rate is modulated in response to stress, metabolic fluxes and developmental status. However, how this control is exerted by the host nucleus is unclear. Here, we report a genome-wide microarray analysis of three accumulation and replication of chloroplasts (arc) mutants that show a spectrum of altered plastid division characteristics. To ensure a comprehensive data set, we selected arc3, arc5 and arc11 because they harbour mutations in protein components of both the stromal and cytosolic division machinery, are of different evolutionary origin and display different phenotypic severities in terms of chloroplast number, size and volume. We show that a surprisingly low number of genes are affected by altered plastid division status, but that the affected genes encode proteins important for a variety of fundamental plant processes.
Asunto(s)
Arabidopsis/genética , División Celular , Plastidios/genética , Transcriptoma , Algoritmos , Arabidopsis/citología , Arabidopsis/metabolismo , Arabidopsis/fisiología , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Citosol/metabolismo , Dinaminas/genética , Dinaminas/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Mutación , Análisis de Secuencia por Matrices de Oligonucleótidos , Fenotipo , Plastidios/metabolismo , Plastidios/fisiología , Estrés FisiológicoRESUMEN
The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of human COVID-19, not only causes flu-like symptoms and gut microbiome complications but a large number of infected individuals also experience a host of neurological symptoms including loss of smell and taste, seizures, difficulty concentrating, decreased alertness, and brain inflammation. Although SARS-CoV-2 infections are not more prevalent in Parkinson's disease patients, a higher mortality rate has been reported not only associated with older age and longer disease duration, but also through several mechanisms, such as interactions with the brain dopaminergic system and through systemic inflammatory responses. Indeed, a number of the neurological symptoms seen in COVID-19 patients, as well as the alterations in the gut microbiome, are also prevalent in patients with Parkinson's disease. Furthermore, biochemical pathways such as oxidative stress, inflammation, and protein aggregation have shared commonalities between Parkinson's disease and COVID-19 disease progression. In this review, we describe and compare the numerous similarities and intersections between neurodegeneration in Parkinson's disease and RNA viral infections, emphasizing the current SARS-CoV-2 global health crisis.
Asunto(s)
COVID-19/fisiopatología , Microbioma Gastrointestinal , Enfermedad de Parkinson/fisiopatología , SARS-CoV-2 , Anciano , COVID-19/complicaciones , COVID-19/mortalidad , Trastornos del Conocimiento/etiología , Citocinas/fisiología , Dieta , Progresión de la Enfermedad , Disbiosis/etiología , Disbiosis/fisiopatología , Humanos , Inflamación , Metales Pesados/toxicidad , Modelos Neurológicos , Degeneración Nerviosa , Bulbo Olfatorio/fisiopatología , Bulbo Olfatorio/virología , Estrés Oxidativo , Enfermedad de Parkinson/etiología , Guías de Práctica Clínica como Asunto , Agregación Patológica de Proteínas/etiología , Infecciones por Virus ARN/metabolismo , Infecciones por Virus ARN/fisiopatología , Especies Reactivas de Oxígeno/metabolismo , Trastornos de la Sensación/etiología , alfa-Sinucleína/metabolismoRESUMEN
70 kDa heat shock proteins (Hsp70s) act as molecular chaperones involved in essential cellular processes such as protein folding and protein transport across membranes. They also play a role in the cell's response to a wide range of stress conditions. The Arabidopsis family of Hsp70s homologues includes two highly conserved proteins, cpHsc70-1 and cpHsc70-2 which are both imported into chloroplasts (Su and Li in Plant Physiol 146:1231-1241, 2008). Here, we demonstrate that YFP-fusion proteins of both cpHsc70-1 and cpHsc70-2 are predominantly stromal, though low levels were detected in the thylakoid membrane. Both genes are ubiquitously expressed at high levels in both seedlings and adult plants. We further show that both cpHsc70-1 and cpHsc70-2 harbour ATPase activity which is essential for Hsp70 chaperone activity. A previously described T-DNA insertion line for cpHsc70-1 (DeltacpHsc70-1) has variegated cotyledons, malformed leaves, growth retardation, impaired root growth and sensitivity to heat shock treatment. In addition, under stress conditions, this mutant also exhibits unusual sepals, and malformed flowers and sucrose concentrations as low as 1% significantly impair growth. cpHsc70-1/cpHsc70-2 double-mutants are lethal. However, we demonstrate through co-suppression and artificial microRNA (amiRNA) approaches that transgenic plants with severely reduced levels of both genes have a white and stunted phenotype. Interestingly, chloroplasts in these plants have an unusual morphology and contain few or no thylakoid membranes. Our data show that cpHsc70-1 and cpHsc70-2 are essential ATPases, have overlapping roles and are required for normal plastid structure.
Asunto(s)
Arabidopsis/metabolismo , Cloroplastos/metabolismo , Proteínas HSP70 de Choque Térmico/metabolismo , Adenosina Trifosfatasas/metabolismo , Arabidopsis/enzimología , Arabidopsis/crecimiento & desarrollo , Western Blotting , Cloroplastos/enzimología , Cartilla de ADN , Silenciador del Gen , Genes Reporteros , MicroARNs/genética , Microscopía Fluorescente , Fotosíntesis , Interferencia de ARNRESUMEN
Plastids are vital organelles, fulfilling important metabolic functions that greatly influence plant growth and productivity. In order to both regulate and harness the metabolic output of plastids, it is vital that the process of plastid division is carefully controlled. This is essential, not only to ensure persistence in dividing plant cells and that optimal numbers of plastids are obtained in specialized cell types, but also to allow the cell to act in response to developmental signals and environmental changes. How this control is exerted by the host nucleus has remained elusive. Plastids evolved by endosymbiosis and during the establishment of a permanent endosymbiosis they retained elements of the bacterial cell-division machinery. Through evolution the photosynthetic eukaryotes have increased dramatically in complexity, from single-cell green algae to multicellular non-vascular and vascular plants. Reflected with this is an increasing complexity of the division machinery and recent findings also suggest increasing complexity in the molecular mechanisms used by the host cell to control the process of plastid division. In the present paper, we explore the current understanding of the process of plastid division at the molecular and cellular level, with particular respect to the evolution of the division machinery and levels of control exerted on the process.