RESUMEN

PURPOSE: Interneuronopathies are a group of neurodevelopmental disorders characterized by deficient migration and differentiation of gamma-aminobutyric acidergic interneurons resulting in a broad clinical spectrum, including autism spectrum disorders, early-onset epileptic encephalopathy, intellectual disability, and schizophrenic disorders. SP9 is a transcription factor belonging to the Krüppel-like factor and specificity protein family, the members of which harbor highly conserved DNA-binding domains. SP9 plays a central role in interneuron development and tangential migration, but it has not yet been implicated in a human neurodevelopmental disorder. METHODS: Cases with SP9 variants were collected through international data-sharing networks. To address the specific impact of SP9 variants, in silico and in vitro assays were carried out. RESULTS: De novo heterozygous variants in SP9 cause a novel form of interneuronopathy. SP9 missense variants affecting the glutamate 378 amino acid result in severe epileptic encephalopathy because of hypomorphic and neomorphic DNA-binding effects, whereas SP9 loss-of-function variants result in a milder phenotype with epilepsy, developmental delay, and autism spectrum disorder. CONCLUSION: De novo heterozygous SP9 variants are responsible for a neurodevelopmental disease. Interestingly, variants located in conserved DNA-binding domains of KLF/SP family transcription factors may lead to neomorphic DNA-binding functions resulting in a combination of loss- and gain-of-function effects.

Asunto(s)

Trastorno del Espectro Autista , Epilepsia , Discapacidad Intelectual , Interneuronas , Factores de Transcripción Sp , Factores de Transcripción , Adolescente , Niño , Preescolar , Femenino , Humanos , Masculino , Trastorno del Espectro Autista/genética , Trastorno del Espectro Autista/patología , Epilepsia/genética , Epilepsia/patología , Heterocigoto , Discapacidad Intelectual/genética , Discapacidad Intelectual/patología , Interneuronas/metabolismo , Interneuronas/patología , Mutación Missense/genética , Trastornos del Neurodesarrollo/genética , Trastornos del Neurodesarrollo/patología , Fenotipo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Factores de Transcripción Sp/genética

RESUMEN

Mendel's law of segregation states that the two alleles at a diploid locus should be transmitted equally to the progeny. A genetic segregation distortion, also referred to as transmission ratio distortion (TRD), is a statistically significant deviation from this rule. TRD has been observed in several mammal species and may be due to different biological mechanisms occurring at diverse time points ranging from gamete formation to lethality at post-natal stages. In this review, we describe examples of TRD and their possible mechanisms in mammals based on current knowledge. We first focus on the differences between TRD in male and female gametogenesis in the house mouse, in which some of the most well studied TRD systems have been characterized. We then describe known TRD in other mammals, with a special focus on the farmed species and in the peculiar common shrew species. Finally, we discuss TRD in human diseases. Thus far, to our knowledge, this is the first time that such description is proposed. This review will help better comprehend the processes involved in TRD. A better understanding of these molecular mechanisms will imply a better comprehension of their impact on fertility and on genome evolution. In turn, this should allow for better genetic counseling and lead to better care for human families.

Asunto(s)

Células Germinativas , Mamíferos , Animales , Ratones , Humanos , Masculino , Femenino , Mamíferos/genética

RESUMEN

CBS encodes a pyridoxal 5'-phosphate-dependent enzyme that catalyses the condensation of homocysteine and serine to form cystathionine. Due to its implication in some cancers and in the cognitive pathophysiology of Down syndrome, the identification of pharmacological inhibitors of this enzyme is urgently required. However, thus far, attempts to identify such molecules have only led to the identification of compounds with low potency and limited selectivity. We consequently developed an original, yeast-based screening method that identified three FDA-approved drugs of the 8-hydroxyquinoline family: clioquinol, chloroxine and nitroxoline. These molecules reduce CBS enzymatic activity in different cellular models, proving that the molecular mechanisms involved in yeast phenotypic rescue are conserved in mammalian cells. A combination of genetic and chemical biology approaches also revealed the importance of copper and zinc intracellular levels in the regulation of CBS enzymatic activity-copper promoting CBS activity and zinc inhibiting its activity. Taken together, these results indicate that our effective screening approach identified three new potent CBS inhibitors and provides new findings for the regulation of CBS activity, which is crucial to develop new therapies for CBS-related human disorders.

Asunto(s)

Cistationina betasintasa , Saccharomyces cerevisiae , Animales , Cobre , Cistationina betasintasa/genética , Humanos , Mamíferos , Oxiquinolina/farmacología , Fosfato de Piridoxal , Zinc

RESUMEN

Prenatal alcohol exposure is a major cause of neurobehavioral disabilities. MRI studies in humans have shown that alcohol is associated with white matter microstructural anomalies but these studies focused on myelin abnormalities only after birth. Only one of these studies evaluated oligodendrocyte lineage, but only for a short period during human foetal life. As data are lacking in humans and alcohol is known to impair oligodendrocyte differentiation in rodents, the present study aimed to compare by immunohistochemistry the oligodendrocyte precursor cells expressing PDGFR-α and immature premyelinating/mature oligodendrocytes expressing Olig2 in the ganglionic eminences and the frontal cortex of 14 human foetuses exposed to alcohol from 15 to 37 weeks' gestation with age-matched controls. The human brains used in this study were obtained at the time of foetal autopsies for medical termination of pregnancy, in utero or post-natal early death. Before birth, PDGFR-α expression was strongly increased in the ganglionic eminences and the cortex of all foetuses exposed to alcohol except at the earliest stage. No massive generation of Olig2 immunoreactive cells was identified in the ganglionic eminences until the end of pregnancy and the density of Olig2-positive cells within the cortex was consistently lower in foetuses exposed to alcohol than in controls. These antenatal data from humans provides further evidence of major oligodendrocyte lineage impairment at specific and key stages of brain development upon prenatal alcohol exposure including defective or delayed generation and maturation of oligodendrocyte precursors.

Asunto(s)

Efectos Tardíos de la Exposición Prenatal , Diferenciación Celular , Linaje de la Célula , Etanol/toxicidad , Femenino , Feto/metabolismo , Humanos , Vaina de Mielina/metabolismo , Factor de Transcripción 2 de los Oligodendrocitos/metabolismo , Oligodendroglía/metabolismo , Embarazo , Efectos Tardíos de la Exposición Prenatal/metabolismo

RESUMEN

Mitochondrial complex V plays an important role in oxidative phosphorylation by catalyzing the generation of ATP. Most complex V subunits are nuclear encoded and not yet associated with recognized Mendelian disorders. Using exome sequencing, we identified a rare homozygous splice variant (c.87+3A>G) in ATP5PO, the complex V subunit which encodes the oligomycin sensitivity conferring protein, in three individuals from two unrelated families, with clinical suspicion of a mitochondrial disorder. These individuals had a similar, severe infantile and often lethal multi-systemic disorder that included hypotonia, developmental delay, hypertrophic cardiomyopathy, progressive epileptic encephalopathy, progressive cerebral atrophy, and white matter abnormalities on brain MRI consistent with Leigh syndrome. cDNA studies showed a predominant shortened transcript with skipping of exon 2 and low levels of the normal full-length transcript. Fibroblasts from the affected individuals demonstrated decreased ATP5PO protein, defective assembly of complex V with markedly reduced amounts of peripheral stalk proteins, and complex V hydrolytic activity. Further, expression of human ATP5PO cDNA without exon 2 (hATP5PO-∆ex2) in yeast cells deleted for yATP5 (ATP5PO homolog) was unable to rescue growth on media which requires oxidative phosphorylation when compared to the wild type construct (hATP5PO-WT), indicating that exon 2 deletion leads to a non-functional protein. Collectively, our findings support the pathogenicity of the ATP5PO c.87+3A>G variant, which significantly reduces but does not eliminate complex V activity. These data along with the recent report of an affected individual with ATP5PO variants, add to the evidence that rare biallelic variants in ATP5PO result in defective complex V assembly, function and are associated with Leigh syndrome.

Asunto(s)

Encefalopatías , Enfermedad de Leigh , ATPasas de Translocación de Protón Mitocondriales , Encefalopatías/metabolismo , ADN Complementario/metabolismo , Humanos , Enfermedad de Leigh/genética , Enfermedad de Leigh/metabolismo , Mitocondrias/genética , Mitocondrias/metabolismo , ATPasas de Translocación de Protón Mitocondriales/genética , Mutación , Proteínas/metabolismo

RESUMEN

Down syndrome (DS), the most frequent chromosomic aberration, results from the presence of an extra copy of chromosome 21. The identification of genes which overexpression contributes to intellectual disability (ID) in DS is important to understand the pathophysiological mechanisms involved and develop new pharmacological therapies. In particular, gene dosage of Dual specificity tyrosine phosphorylation Regulated Kinase 1A (DYRK1A) and of Cystathionine beta synthase (CBS) are crucial for cognitive function. As these two enzymes have lately been the main targets for therapeutic research on ID, we sought to decipher the genetic relationship between them. We also used a combination of genetic and drug screenings using a cellular model overexpressing CYS4, the homolog of CBS in Saccharomyces cerevisiae, to get further insights into the molecular mechanisms involved in the regulation of CBS activity. We showed that overexpression of YAK1, the homolog of DYRK1A in yeast, increased CYS4 activity whereas GSK3ß was identified as a genetic suppressor of CBS. In addition, analysis of the signaling pathways targeted by the drugs identified through the yeast-based pharmacological screening, and confirmed using human HepG2 cells, emphasized the importance of Akt/GSK3ß and NF-κB pathways into the regulation of CBS activity and expression. Taken together, these data provide further understanding into the regulation of CBS and in particular into the genetic relationship between DYRK1A and CBS through the Akt/GSK3ß and NF-κB pathways, which should help develop more effective therapies to reduce cognitive deficits in people with DS.

RESUMEN

Prion diseases are caused by the propagation of PrPSc, the pathological conformation of the PrPC prion protein. The molecular mechanisms underlying PrPSc propagation are still unsolved and no therapeutic solution is currently available. We thus sought to identify new anti-prion molecules and found that flunarizine inhibited PrPSc propagation in cell culture and significantly prolonged survival of prion-infected mice. Using an in silico therapeutic repositioning approach based on similarities with flunarizine chemical structure, we tested azelastine, duloxetine, ebastine, loperamide and metixene and showed that they all have an anti-prion activity. Like flunarizine, these marketed drugs reduced PrPSc propagation in cell culture and in mouse cerebellum organotypic slice culture, and inhibited the protein folding activity of the ribosome (PFAR). Strikingly, some of these drugs were also able to alleviate phenotypes due to PABPN1 nuclear aggregation in cell and Drosophila models of oculopharyngeal muscular dystrophy (OPMD). These data emphasize the therapeutic potential of anti-PFAR drugs for neurodegenerative and neuromuscular proteinopathies.

Asunto(s)

Sistemas de Liberación de Medicamentos/métodos , Flunarizina/administración & dosificación , Proteína I de Unión a Poli(A)/metabolismo , Enfermedades por Prión/metabolismo , Agregado de Proteínas/efectos de los fármacos , Pliegue de Proteína/efectos de los fármacos , Animales , Bloqueadores de los Canales de Calcio/administración & dosificación , Línea Celular , Bases de Datos Factuales , Drosophila , Femenino , Ratones , Ratones Transgénicos , Técnicas de Cultivo de Órganos , Proteína I de Unión a Poli(A)/antagonistas & inhibidores , Proteína I de Unión a Poli(A)/genética , Enfermedades por Prión/tratamiento farmacológico , Enfermedades por Prión/genética , Proteínas Priónicas/antagonistas & inhibidores , Proteínas Priónicas/genética , Proteínas Priónicas/metabolismo , Agregado de Proteínas/fisiología , Ovinos

RESUMEN

Identified in the late 1970s as an oncogene, a driving force leading to tumor development, p53 turned out to be a key tumor suppressor gene. Now p53 is considered a master gene regulating the transcription of over 3000 target genes and controlling a remarkable number of cellular functions. The elevated prevalence of p53 mutations in human cancers has led to a recurring questioning about the roles of mutant p53 proteins and their functional consequences. Both mutants and isoforms of p53 have been attributed dominant-negative and gain of function properties among which is the ability to form amyloid aggregates and behave in a prion-like manner. This report challenges the ongoing "prion p53" hypothesis by reviewing evidence of p53 behavior in light of our current knowledge regarding amyloid proteins, prionoids and prions.

RESUMEN

Alcohol affects multiple neurotransmitter systems, notably the GABAergic system and has been recognised for a long time as particularly damaging during critical stages of brain development. Nevertheless, data from the literature are most often derived from animal or in vitro models. In order to study the production, migration and cortical density disturbances of GABAergic interneurons upon prenatal alcohol exposure, we performed immunohistochemical studies by means of the proliferation marker Ki67, GABA and calretinin antibodies in the frontal cortical plate of 17 foetal and infant brains antenatally exposed to alcohol, aged 15 weeks' gestation to 22 postnatal months and in the ganglionic eminences and the subventricular zone of the dorsal telencephalon until their regression, i.e., 34 weeks' gestation. Results were compared with those obtained in 17 control brains aged 14 weeks of gestation to 35 postnatal months. We also focused on interneuron vascular migration along the cortical microvessels by confocal microscopy with double immunolabellings using Glut1, GABA and calretinin. Semi-quantitative and quantitative analyses of GABAergic and calretininergic interneuron density allowed us to identify an insufficient and delayed production of GABAergic interneurons in the ganglionic eminences during the two first trimesters of the pregnancy and a delayed incorporation into the laminar structures of the frontal cortex. Moreover, a mispositioning of GABAergic and calretininergic interneurons persisted throughout the foetal life, these cells being located in the deep layers instead of the superficial layers II and III. Moreover, vascular migration of calretininergic interneurons within the cortical plate was impaired, as reflected by low numbers of interneurons observed close to the cortical perforating vessel walls that may in part explain their abnormal intracortical distribution. Our results are globally concordant with those previously obtained in mouse models, in which alcohol has been shown to induce an interneuronopathy by affecting interneuron density and positioning within the cortical plate, and which could account for the neurological disabilities observed in children with foetal alcohol disorder spectrum.

Asunto(s)

Consumo de Bebidas Alcohólicas , Encéfalo/embriología , Calbindina 2/metabolismo , Trastornos del Espectro Alcohólico Fetal/metabolismo , Feto/embriología , Interneuronas/metabolismo , Antígeno Ki-67/metabolismo , Efectos Tardíos de la Exposición Prenatal/metabolismo , Ácido gamma-Aminobutírico/metabolismo , Alcoholismo , Consumo Excesivo de Bebidas Alcohólicas , Encéfalo/metabolismo , Encéfalo/patología , Estudios de Casos y Controles , Movimiento Celular , Femenino , Trastornos del Espectro Alcohólico Fetal/patología , Feto/metabolismo , Feto/patología , Lóbulo Frontal/embriología , Lóbulo Frontal/metabolismo , Lóbulo Frontal/patología , Neuronas GABAérgicas/metabolismo , Neuronas GABAérgicas/patología , Humanos , Lactante , Recién Nacido , Interneuronas/patología , Masculino , Embarazo , Complicaciones del Embarazo , Segundo Trimestre del Embarazo , Efectos Tardíos de la Exposición Prenatal/patología , Telencéfalo/embriología , Telencéfalo/metabolismo , Telencéfalo/patología

RESUMEN

BACKGROUND: Hydrogen sulfide (H2S) is an endogenous mammalian gasotransmitter. Cystathionine ß-synthase (CBS), cystathionine γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (3-MST) are the principal enzymes responsible for its biogenesis. A recent yeast screen suggested that disulfiram (a well-known inhibitor of aldehyde dehydrogenase and a clinically used drug in the treatment of alcoholism) may inhibit CBS in a cell-based environment. However, prior studies have not observed any direct inhibition of CBS by disulfiram. We investigated the potential role of bioconversion of disulfiram to bis(N,N-diethyldithiocarbamate)-copper(II) complex (CuDDC) in the inhibitory effect of disulfiram on H2S production and assessed its effect in two human cell types with high CBS expression: HCT116 colon cancer cells and Down syndrome (DS) fibroblasts. METHODS: H2S production from recombinant human CBS, CSE and 3-MST was measured using the fluorescent H2S probe AzMC. Mouse liver homogenate (a rich source of CBS) was also employed to measure H2S biosynthesis. The interaction of copper with accessible protein cysteine residues was evaluated using the DTNB method. Cell proliferation and viability were measured using the BrdU and MTT methods. Cellular bioenergetics was evaluated by Extracellular Flux Analysis. RESULTS: While disulfiram did not exert any significant direct inhibitory effect on any of the H2S-producing enzymes, its metabolite, CuDDC was a potent inhibitor of CBS and CSE. The mode of its action is likely related to the complexed copper molecule. In cell-based systems, the effects of disulfiram were variable. In colon cancer cells, no significant effect of disulfiram was observed on H2S production or proliferation or viability. In contrast, in DS fibroblasts, disulfiram inhibited H2S production and improved proliferation and viability. Copper, on its own, failed to have any effects on either cell type, likely due to its low cell penetration. CuDDC inhibited H2S production in both cell types studied and exerted the functional effects that would be expected from a CBS inhibitor: inhibition of cell proliferation of cancer cells and a bell-shaped effect (stimulation of proliferation at low concentration and inhibition of these responses at higher concentration) in DS cells. Control experiments using a chemical H2S donor showed that, in addition to inhibiting CBS and CSE, part of the biological effects of CuDDC relates to a direct reaction with H2S, which occurs through its complexed copper. CONCLUSIONS: Disulfiram, via its metabolite CuDDC acts as an inhibitor of CBS and a scavenger of H2S, which, in turn, potently suppresses H2S levels in various cell types. Inhibition of H2S biosynthesis may explain some of the previously reported actions of disulfiram and CuDDC in vitro and in vivo. Disulfiram or CuDDC may be considered as potential agents for the experimental therapy of various pathophysiological conditions associated with H2S overproduction.

Asunto(s)

Inhibidores del Acetaldehído Deshidrogenasa/farmacología , Cobre/farmacología , Cistationina betasintasa/antagonistas & inhibidores , Disulfiram/farmacología , Ditiocarba/análogos & derivados , Compuestos Organometálicos/farmacología , Inhibidores del Acetaldehído Deshidrogenasa/metabolismo , Animales , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Quelantes/metabolismo , Quelantes/farmacología , Cobre/metabolismo , Cistationina betasintasa/metabolismo , Disulfiram/metabolismo , Ditiocarba/metabolismo , Ditiocarba/farmacología , Relación Dosis-Respuesta a Droga , Femenino , Células HCT116 , Humanos , Hígado/efectos de los fármacos , Hígado/metabolismo , Ratones , Ratones Endogámicos BALB C , Compuestos Organometálicos/metabolismo

RESUMEN

Mitochondria continually move, fuse and divide, and these dynamics are essential for the proper function of the organelles. Indeed, the dynamic balance of fusion and fission of mitochondria determines their morphology and allows their immediate adaptation to energetic needs as well as preserving their integrity. As a consequence, mitochondrial fusion and fission dynamics and the proteins that control these processes, which are conserved from yeast to human, are essential, and their disturbances are associated with severe human disorders, including neurodegenerative diseases. For example, mutations in OPA1, which encodes a conserved factor essential for mitochondrial fusion, lead to optic atrophy 1, a neurodegeneration that affects the optic nerve, eventually leading to blindness. Here, by screening a collection of ∼1600 repurposed drugs on a fission yeast model, we identified five compounds able to efficiently prevent the lethality associated with the loss of Msp1p, the fission yeast ortholog of OPA1. One compound, hexestrol, was able to rescue both the mitochondrial fragmentation and mitochondrial DNA (mtDNA) depletion induced by the loss of Msp1p, whereas the second, clomifene, only suppressed the mtDNA defect. Yeast has already been successfully used to identify candidate drugs to treat inherited mitochondrial diseases; this work may therefore provide useful leads for the treatment of optic atrophies such as optic atrophy 1 or Leber hereditary optic neuropathy.

Asunto(s)

ADN Mitocondrial/metabolismo , Evaluación Preclínica de Medicamentos , Reposicionamiento de Medicamentos , Dinámicas Mitocondriales , Schizosaccharomyces/metabolismo , Clomifeno/farmacología , Hexestrol/farmacología , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Dinámicas Mitocondriales/efectos de los fármacos , Dominios Proteicos , Proteínas de Schizosaccharomyces pombe/química , Proteínas de Schizosaccharomyces pombe/metabolismo

RESUMEN

Identifying dosage-sensitive genes is a key to understand the mechanisms underlying intellectual disability in Down syndrome (DS). The Dp(17Abcg1-Cbs)1Yah DS mouse model (Dp1Yah) shows cognitive phenotypes that need to be investigated to identify the main genetic driver. Here, we report that three copies of the cystathionine-beta-synthase gene (Cbs) in the Dp1Yah mice are necessary to observe a deficit in the novel object recognition (NOR) paradigm. Moreover, the overexpression of Cbs alone is sufficient to induce deficits in the NOR test. Accordingly, overexpressing human CBS specifically in Camk2a-expressing neurons leads to impaired objects discrimination. Altogether, this shows that Cbs overdosage is involved in DS learning and memory phenotypes. To go further, we identified compounds that interfere with the phenotypical consequence of CBS overdosage in yeast. Pharmacological intervention in Tg(CBS) mice with one selected compound restored memory in the NOR test. In addition, using a genetic approach, we demonstrated an epistatic interaction between Cbs and Dyrk1a, another human chromosome 21-located gene (which encodes the dual-specificity tyrosine phosphorylation-regulated kinase 1a) and an already known target for DS therapeutic intervention. Further analysis using proteomic approaches highlighted several molecular pathways, including synaptic transmission, cell projection morphogenesis and actin cytoskeleton, that are affected by DYRK1A and CBS overexpression. Overall, we demonstrated that CBS overdosage underpins the DS-related recognition memory deficit and that both CBS and DYRK1A interact to control accurate memory processes in DS. In addition, our study establishes CBS as an intervention point for treating intellectual deficiencies linked to DS.

Asunto(s)

Cistationina betasintasa/genética , Síndrome de Down/diagnóstico , Síndrome de Down/genética , Epistasis Genética , Dosificación de Gen , Fenotipo , Proteínas Serina-Treonina Quinasas/genética , Proteínas Tirosina Quinasas/genética , Animales , Conducta Animal , Cognición , Modelos Animales de Enfermedad , Humanos , Locomoción , Memoria , Ratones , Ratones Transgénicos , Neuronas/metabolismo , Proteoma , Proteómica/métodos , Quinasas DyrK

RESUMEN

The ARX (Aristaless Related homeoboX) gene was identified in 2002 as responsible for XLAG syndrome, a lissencephaly characterized by an almost complete absence of cortical GABAergic interneurons, and for milder forms of X-linked Intellectual Disability (ID) without apparent brain abnormalities. The most frequent mutation found in the ARX gene, a duplication of 24 base pairs (c.429_452dup24) in exon 2, results in a recognizable syndrome in which patients present ID without primary motor impairment, but with a very specific upper limb distal motor apraxia associated with a pathognomonic hand-grip, described as developmental Limb Kinetic Apraxia (LKA). In this study, we first present ARX expression during human fetal brain development showing that it is strongly expressed in GABAergic neuronal progenitors during the second and third trimester of pregnancy. We show that although ARX expression strongly decreases towards the end of gestation, it is still present after birth in some neurons of the basal ganglia, thalamus and cerebral cortex, suggesting that ARX also plays a role in more mature neuron functioning. Then, using morphometric brain MRI in 13 ARX patients carrying c.429_452dup24 mutation and in 13 sex- and age-matched healthy controls, we show that ARX patients have a significantly decreased volume of several brain structures including the striatum (and more specifically the caudate nucleus), hippocampus and thalamus as well as decreased precentral gyrus cortical thickness. We observe a significant correlation between caudate nucleus volume reduction and motor impairment severity quantified by kinematic parameter of precision grip. As basal ganglia are known to regulate sensorimotor processing and are involved in the control of precision gripping, the combined decrease in cortical thickness of primary motor cortex and basal ganglia volume in ARX dup24 patients is very likely the anatomical substrate of this developmental form of LKA.

Asunto(s)

Ganglios Basales/metabolismo , Genes Homeobox/genética , Proteínas de Homeodominio/genética , Mutación/genética , Factores de Transcripción/genética , Apraxia Ideomotora/genética , Proteína Doblecortina , Femenino , Fuerza de la Mano/fisiología , Humanos , Interneuronas/metabolismo , Neuronas/metabolismo , Embarazo , Ácido gamma-Aminobutírico/metabolismo

RESUMEN

The aristaless-related homeobox (ARX) transcription factor is involved in the development of GABAergic and cholinergic neurons in the forebrain. ARX mutations have been associated with a wide spectrum of neurodevelopmental disorders in humans, among which the most frequent, a 24 bp duplication in the polyalanine tract 2 (c.428_451dup24), gives rise to intellectual disability, fine motor defects with or without epilepsy. To understand the functional consequences of this mutation, we generated a partially humanized mouse model carrying the c.428_451dup24 duplication (Arxdup24/0) that we characterized at the behavior, neurological and molecular level. Arxdup24/0 males presented with hyperactivity, enhanced stereotypies and altered contextual fear memory. In addition, Arxdup24/0 males had fine motor defects with alteration of reaching and grasping abilities. Transcriptome analysis of Arxdup24/0 forebrains at E15.5 showed a down-regulation of genes specific to interneurons and an up-regulation of genes normally not expressed in this cell type, suggesting abnormal interneuron development. Accordingly, interneuron migration was altered in the cortex and striatum between E15.5 and P0 with consequences in adults, illustrated by the defect in the inhibitory/excitatory balance in Arxdup24/0 basolateral amygdala. Altogether, we showed that the c.428_451dup24 mutation disrupts Arx function with a direct consequence on interneuron development, leading to hyperactivity and defects in precise motor movement control and associative memory. Interestingly, we highlighted striking similarities between the mouse phenotype and a cohort of 33 male patients with ARX c.428_451dup24, suggesting that this new mutant mouse line is a good model for understanding the pathophysiology and evaluation of treatment.

Asunto(s)

Epilepsia/genética , Proteínas de Homeodominio/genética , Trastornos del Neurodesarrollo/genética , Factores de Transcripción/genética , Adolescente , Adulto , Animales , Niño , Preescolar , Neuronas Colinérgicas/metabolismo , Neuronas Colinérgicas/patología , Contractura , Modelos Animales de Enfermedad , Epilepsia/fisiopatología , Neuronas GABAérgicas/metabolismo , Neuronas GABAérgicas/patología , Regulación del Desarrollo de la Expresión Génica , Humanos , Lactante , Discapacidad Intelectual , Masculino , Ratones , Mutación , Trastornos del Neurodesarrollo/fisiopatología , Péptidos/genética , Prosencéfalo/fisiopatología , Paraplejía Espástica Hereditaria , Transcriptoma/genética , Adulto Joven

RESUMEN

It is no longer necessary to demonstrate that ribosome is the central machinery of protein synthesis. But it is less known that it is also key player of the protein folding process through another conserved function: the protein folding activity of the ribosome (PFAR). This ribozyme activity, discovered more than 2 decades ago, depends upon the domain V of the large rRNA within the large subunit of the ribosome. Surprisingly, we discovered that anti-prion compounds are also potent PFAR inhibitors, highlighting an unexpected link between PFAR and prion propagation. In this review, we discuss the ancestral origin of PFAR in the light of the ancient RNA world hypothesis. We also consider how this ribosomal activity fits into the landscape of cellular protein chaperones involved in the appearance and propagation of prions and other amyloids in mammals. Finally, we examine how drugs targeting the protein folding activity of the ribosome could be active against mammalian prion and other protein aggregation-based diseases, making PFAR a promising therapeutic target for various human protein misfolding diseases.

Asunto(s)

Priones/metabolismo , Pliegue de Proteína , Ribosomas/metabolismo , Ribosomas/patología , Animales , Proteínas de Choque Térmico/metabolismo , Humanos , Modelos Moleculares , Enfermedades por Prión/metabolismo , Enfermedades por Prión/patología , Priones/química , Biosíntesis de Proteínas , ARN Ribosómico/metabolismo

RESUMEN

The Aristaless-related homeobox (ARX) gene encodes a paired-type homeodomain transcription factor with critical roles in embryonic development. Mutations in ARX give rise to intellectual disability (ID), epilepsy and brain malformation syndromes. To capture the genetics and molecular disruptions that underpin the ARX-associated clinical phenotypes, we undertook a transcriptome wide RNASeq approach to analyse developing (12.5 dpc) telencephalon of mice modelling two recurrent polyalanine expansion mutations with different phenotypic severities in the ARX gene. Here we report 238 genes significantly deregulated (Log2FC > +/-1.1, P-value <0.05) when both mutations are compared to wild-type (WT) animals. When each mutation is considered separately, a greater number of genes were deregulated in the severe PA1 mice (825) than in the PA2 animals (78). Analysing genes deregulated in either or both mutant strains, we identified 12% as implicated in ID, epilepsy and autism (99/858), with ∼5% of them as putative or known direct targets of ARX transcriptional regulation. We propose a core pathway of transcription regulators, including Hdac4, involved in chromatin condensation and transcriptional repression, and one of its targets, the transcription factor Twist1, as potential drivers of the ID and infantile spasms in patients with ARX polyalanine expansion mutations. We predict that the subsequent disturbance to this pathway is a consequence of ARX protein reduction with a broader and more significant level of disruption in the PA1 in comparison to the PA2 mice. Identifying early triggers of ARX-associated phenotypes contributes to our understanding of particular clusters/pathways underpinning comorbid phenotypes that are shared by many neurodevelopmental disorders.

Asunto(s)

Epilepsia/genética , Proteínas de Homeodominio/genética , Discapacidad Intelectual/genética , Péptidos/genética , Factores de Transcripción/genética , Transcriptoma/genética , Animales , Modelos Animales de Enfermedad , Epilepsia/patología , Regulación del Desarrollo de la Expresión Génica , Histona Desacetilasas/genética , Humanos , Discapacidad Intelectual/patología , Ratones , Mutación , Fenotipo , Prosencéfalo/embriología , Prosencéfalo/metabolismo , Biosíntesis de Proteínas/genética , Transducción de Señal , Telencéfalo/embriología , Telencéfalo/metabolismo

RESUMEN

The tumor suppression activity of p53 is frequently impaired in cancers even when a wild-type copy of the gene is still present, suggesting that a dominant-negative effect is exerted by some of p53 mutants and isoforms. p63 and p73, which are related to p53, have also been reported to be subjected to a similar loss of function, suggesting that a dominant-negative interplay might happen between p53, p63 and p73. However, to which extent p53 hotspot mutants and isoforms of p53, p63 and p73 are able to interfere with the tumor suppressive activity of their siblings as well as the underlying mechanisms remain undeciphered. Using yeast, we showed that a dominant-negative effect is widely spread within the p53/p63/p73 family as all p53 loss-of-function hotspot mutants and several of the isoforms of p53 and p73 tested exhibit a dominant-negative potential. In addition, we found that this dominant-negative effect over p53 wild-type is based on tetramer poisoning through the formation of inactive hetero-tetramers and does not rely on a prion-like mechanism contrary to what has been previously suggested. We also showed that mutant p53-R175H gains the ability to inhibit p63 and p73 activity by a mechanism that is only partially based on tetramerization.

Asunto(s)

Regulación de la Expresión Génica , Factores de Transcripción/genética , Proteína Tumoral p73/genética , Proteína p53 Supresora de Tumor/genética , Proteínas Supresoras de Tumor/genética , Humanos , Mutación , Unión Proteica , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Saccharomyces cerevisiae/genética , Factores de Transcripción/metabolismo , Proteína Tumoral p73/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Proteínas Supresoras de Tumor/metabolismo

RESUMEN

6AP and GA are potent inhibitors of yeast and mammalian prions and also specific inhibitors of PFAR, the protein-folding activity borne by domain V of the large rRNA of the large subunit of the ribosome. We therefore explored the link between PFAR and yeast prion [PSI(+)] using both PFAR-enriched mutants and site-directed methylation. We demonstrate that PFAR is involved in propagation and de novo formation of [PSI(+)]. PFAR and the yeast heat-shock protein Hsp104 partially compensate each other for [PSI(+)] propagation. Our data also provide insight into new functions for the ribosome in basal thermotolerance and heat-shocked protein refolding. PFAR is thus an evolutionarily conserved cell component implicated in the prion life cycle, and we propose that it could be a potential therapeutic target for human protein misfolding diseases.

Asunto(s)

Proteínas de Choque Térmico/metabolismo , Factores de Terminación de Péptidos/metabolismo , Priones/metabolismo , Pliegue de Proteína , Ribosomas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Guanabenzo/farmacología , Proteínas de Choque Térmico/genética , Mutación , Factores de Terminación de Péptidos/genética , Fenantridinas/farmacología , Priones/genética , Pliegue de Proteína/efectos de los fármacos , ARN Ribosómico/metabolismo , Ribosomas/genética , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética

RESUMEN

The Epstein-Barr gammaherpesvirus (EBV) is the first oncogenic virus discovered in human. Indeed, EBV has been known for more than 50 years to be tightly associated with certain human cancers. As such, EBV has been the subject of extensive studies aiming at deciphering various aspects of its biological cycle, ranging from the regulation of its genome replication and maintenance to the induction of its lytic cycle, including the mechanisms that allow its immune evasion or that are related to its tumorogenicity. For more than 30 years the budding yeast Saccharomyces cerevisiae has fruitfully contributed to a number of these studies. The aim of this article is to review the various aspects of EBV biology for which yeast has been instrumental, and to propose new possible applications for these yeast-based assays, as well as the creation of further yeast models dedicated to EBV. This review article illustrates the tremendous potential of S. cerevisiae in integrated chemobiological approaches for the biomedical research.

Asunto(s)

Investigación Biomédica/métodos , Infecciones por Virus de Epstein-Barr , Herpesvirus Humano 4 , Interacciones Huésped-Patógeno , Modelos Inmunológicos , Saccharomyces cerevisiae , Bioensayo , Infecciones por Virus de Epstein-Barr/inmunología , Infecciones por Virus de Epstein-Barr/virología , Genoma Viral , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/inmunología , Herpesvirus Humano 4/patogenicidad , Humanos , Saccharomyces cerevisiae/inmunología , Saccharomyces cerevisiae/virología

RESUMEN

Cystic Fibrosis Transmembrane conductance Regulator (CFTR) protein has recently been shown to be expressed in the human adult central nervous system (CNS). As CFTR expression has also been documented during embryonic development in several organs, such as the respiratory tract, the intestine and the male reproductive system, suggesting a possible role during development we decided to investigate the expression of CFTR in the human developing CNS. In addition, as some, although rare, neurological symptoms have been reported in patients with CF, we compared the expression of normal and mutated CFTR at several fetal stages. Immunohistochemistry was performed on brain and spinal cord samples of foetuses between 13 and 40 weeks of gestation and compared with five patients with cystic fibrosis (CF) of similar ages. We showed in this study that CFTR is only expressed in neurons and has an early and widespread distribution during development. Although we did not observe any cerebral abnormality in patients with CF, we observed a slight delay in the maturation of several brain structures. We also observed different expression and localization of CFTR depending on the brain structure or the cell maturation stage. Our findings, along with a literature review on the neurological phenotypes of patients with CF, suggest that this gene may play previously unsuspected roles in neuronal maturation or function.

Asunto(s)

Encéfalo/embriología , Encéfalo/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Fibrosis Quística/metabolismo , Encéfalo/citología , Fibrosis Quística/patología , Regulador de Conductancia de Transmembrana de Fibrosis Quística/análisis , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Humanos , Inmunohistoquímica , Lactante , Mutación