RESUMEN
Amyotrophic lateral sclerosis (ALS) is a heterogenous neurodegenerative disorder that affects motor neurons and voluntary muscle control1. ALS heterogeneity includes the age of manifestation, the rate of progression and the anatomical sites of symptom onset. Disease-causing mutations in specific genes have been identified and define different subtypes of ALS1. Although several ALS-associated genes have been shown to affect immune functions2, whether specific immune features account for ALS heterogeneity is poorly understood. Amyotrophic lateral sclerosis-4 (ALS4) is characterized by juvenile onset and slow progression3. Patients with ALS4 show motor difficulties by the time that they are in their thirties, and most of them require devices to assist with walking by their fifties. ALS4 is caused by mutations in the senataxin gene (SETX). Here, using Setx knock-in mice that carry the ALS4-causative L389S mutation, we describe an immunological signature that consists of clonally expanded, terminally differentiated effector memory (TEMRA) CD8 T cells in the central nervous system and the blood of knock-in mice. Increased frequencies of antigen-specific CD8 T cells in knock-in mice mirror the progression of motor neuron disease and correlate with anti-glioma immunity. Furthermore, bone marrow transplantation experiments indicate that the immune system has a key role in ALS4 neurodegeneration. In patients with ALS4, clonally expanded TEMRA CD8 T cells circulate in the peripheral blood. Our results provide evidence of an antigen-specific CD8 T cell response in ALS4, which could be used to unravel disease mechanisms and as a potential biomarker of disease state.
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Esclerosis Amiotrófica Lateral , Linfocitos T CD8-positivos , Células Clonales , Esclerosis Amiotrófica Lateral/inmunología , Esclerosis Amiotrófica Lateral/patología , Animales , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/patología , Células Clonales/patología , ADN Helicasas/genética , ADN Helicasas/metabolismo , Técnicas de Sustitución del Gen , Ratones , Neuronas Motoras/patología , Enzimas Multifuncionales/genética , Enzimas Multifuncionales/metabolismo , Mutación , ARN Helicasas/genética , ARN Helicasas/metabolismoRESUMEN
DDX3X syndrome is a neurodevelopmental disorder accounting for up to 3% of cases of intellectual disability (ID) and affecting primarily females. Individuals diagnosed with DDX3X syndrome can also present with behavioral challenges, motor delays and movement disorders, epilepsy, and congenital malformations. DDX3X syndrome is caused by mutations in the X-linked gene DDX3X, which encodes a DEAD-box RNA helicase with critical roles in RNA metabolism, including mRNA translation. Emerging discoveries from animal models are unveiling a fundamental role of DDX3X in neuronal differentiation and development, especially in the neocortex. Here, we review the current knowledge of genetic and neurobiological mechanisms underlying DDX3X syndrome and their relationship with clinical phenotypes.
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Transcriptionally silent chromatin localizes to the nuclear periphery, which provides a special microenvironment for gene repression. A variety of nuclear membrane proteins interact with repressed chromatin, yet the functional role of these interactions remains poorly understood. Here, we show that, in Schizosaccharomyces pombe, the nuclear membrane protein Lem2 associates with chromatin and mediates silencing and heterochromatin localization. Unexpectedly, we found that these functions can be separated and assigned to different structural domains within Lem2, excluding a simple tethering mechanism. Chromatin association and tethering of centromeres to the periphery are mediated by the N-terminal LEM (LAP2-Emerin-MAN1) domain of Lem2, whereas telomere anchoring and heterochromatin silencing require exclusively its conserved C-terminal MSC (MAN1-Src1 C-terminal) domain. Particularly, silencing by Lem2 is epistatic with the Snf2/HDAC (histone deacetylase) repressor complex SHREC at telomeres, while its necessity can be bypassed by deleting Epe1, a JmjC protein with anti-silencing activity. Furthermore, we found that loss of Lem2 reduces heterochromatin association of SHREC, which is accompanied by increased binding of Epe1. This reveals a critical function of Lem2 in coordinating these antagonistic factors at heterochromatin. The distinct silencing and localization functions mediated by Lem2 suggest that these conserved LEM-containing proteins go beyond simple tethering to play active roles in perinuclear silencing.
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Heterocromatina/metabolismo , Proteínas Nucleares/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Eliminación de Gen , Silenciador del Gen , Proteínas Nucleares/genética , Unión Proteica , Estructura Terciaria de Proteína , Transporte de Proteínas/genética , Schizosaccharomyces , Proteínas de Schizosaccharomyces pombe/genéticaRESUMEN
Niemann-Pick type C (NPC) disease is a lysosomal storage disorder characterized by cholesterol accumulation caused by loss-of-function mutations in the Npc1 gene. NPC disease primarily affects the brain, causing neuronal damage and affecting motor coordination. In addition, considerable liver malfunction in NPC disease is common. Recently, we found that the depletion of annexin A6 (ANXA6), which is most abundant in the liver and involved in cholesterol transport, ameliorated cholesterol accumulation in Npc1 mutant cells. To evaluate the potential contribution of ANXA6 in the progression of NPC disease, double-knockout mice (Npc1-/-/Anxa6-/-) were generated and examined for lifespan, neurologic and hepatic functions, as well as liver histology and ultrastructure. Interestingly, lack of ANXA6 in NPC1-deficient animals did not prevent the cerebellar degeneration phenotype, but further deteriorated their compromised hepatic functions and reduced their lifespan. Moreover, livers of Npc1-/-/Anxa6-/- mice contained a significantly elevated number of foam cells congesting the sinusoidal space, a feature commonly associated with inflammation. We hypothesize that ANXA6 deficiency in Npc1-/- mice not only does not reverse neurologic and motor dysfunction, but further worsens overall liver function, exacerbating hepatic failure in NPC disease.
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Anexina A6/fisiología , Péptidos y Proteínas de Señalización Intracelular/fisiología , Hepatopatías/patología , Longevidad , Animales , Conducta Animal , Hepatopatías/etiología , Hepatopatías/metabolismo , Ratones , Ratones Noqueados , Proteína Niemann-Pick C1RESUMEN
Telomeres and the shelterin complex cap and protect the ends of chromosomes. Telomeres are flanked by the subtelomeric sequences that have also been implicated in telomere regulation, although their role is not well defined. Here, we show that, in Schizosaccharomyces pombe, the telomere-associated sequences (TAS) present on most subtelomeres are hyper-recombinogenic, have metastable nucleosomes, and unusual low levels of H3K9 methylation. Ccq1, a subunit of shelterin, protects TAS from nucleosome loss by recruiting the heterochromatic repressor complexes CLRC and SHREC, thereby linking nucleosome stability to gene silencing. Nucleosome instability at TAS is independent of telomeric repeats and can be transmitted to an intrachromosomal locus containing an ectopic TAS fragment, indicating that this is an intrinsic property of the underlying DNA sequence. When telomerase recruitment is compromised in cells lacking Ccq1, DNA sequences present in the TAS promote recombination between chromosomal ends, independent of nucleosome abundance, implying an active function of these sequences in telomere maintenance. We propose that Ccq1 and fragile subtelomeres co-evolved to regulate telomere plasticity by controlling nucleosome occupancy and genome stability.
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Inestabilidad Genómica/genética , Nucleosomas/genética , Proteínas de Schizosaccharomyces pombe/genética , Telómero/genética , Genoma Fúngico/genética , Heterocromatina/genética , Humanos , Metilación , Schizosaccharomyces/genéticaRESUMEN
Huntington's disease is a neurodegenerative disorder caused by a CAG repeat expansion in exon 1 of the huntingtin gene. Striatal projection neurons are mainly affected, leading to motor symptoms, but molecular mechanisms involved in their vulnerability are not fully characterized. Here, we show that eIF4E binding protein (4E-BP), a protein that inhibits translation, is inactivated in Huntington's disease striatum by increased phosphorylation. Accordingly, we detected aberrant de novo protein synthesis. Proteomic characterization indicates that translation specifically affects sets of proteins as we observed upregulation of ribosomal and oxidative phosphorylation proteins and downregulation of proteins related to neuronal structure and function. Interestingly, treatment with the translation inhibitor 4EGI-1 prevented R6/1 mice motor deficits, although corticostriatal long-term depression was not markedly changed in behaving animals. At the molecular level, injection of 4EGI-1 normalized protein synthesis and ribosomal content in R6/1 mouse striatum. In conclusion, our results indicate that dysregulation of protein synthesis is involved in mutant huntingtin-induced striatal neuron dysfunction.
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Factor 4E Eucariótico de Iniciación/fisiología , Enfermedad de Huntington/genética , Biosíntesis de Proteínas/fisiología , Animales , Conducta Animal , Cuerpo Estriado/metabolismo , Modelos Animales de Enfermedad , Factor 4E Eucariótico de Iniciación/genética , Humanos , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Enfermedad de Huntington/metabolismo , Interneuronas/metabolismo , Masculino , Ratones , Ratones Transgénicos , Neostriado/patología , Degeneración Nerviosa/patología , Neuronas/metabolismo , Proteínas Nucleares/genética , Fosforilación , ProteómicaRESUMEN
Huntington's disease (HD) is a hereditary neurodegenerative disorder caused by an expansion of a CAG repeat in the huntingtin (htt) gene, which results in an aberrant form of the protein (mhtt). This leads to motor and cognitive deficits associated with corticostriatal and hippocampal alterations. The levels of STriatal-Enriched protein tyrosine Phosphatase (STEP), a neural-specific tyrosine phosphatase that opposes the development of synaptic strengthening, are decreased in the striatum of HD patients and also in R6/1 mice, thereby contributing to the resistance to excitotoxicity described in this HD mouse model. Here, we aimed to analyze whether STEP inactivation plays a role in the pathophysiology of HD by investigating its effect on motor and cognitive impairment in the R6/1 mouse model of HD. We found that genetic deletion of STEP delayed the onset of motor dysfunction and prevented the appearance of cognitive deficits in R6/1 mice. This phenotype was accompanied by an increase in pERK1/2 levels, a delay in the decrease of striatal DARPP-32 levels and a reduction in the size of mhtt aggregates, both in the striatum and CA1 hippocampal region. We also found that acute pharmacological inhibition of STEP with TC-2153 improved cognitive function in R6/1 mice. In conclusion, our results show that deletion of STEP has a beneficial effect on motor coordination and cognition in a mouse model of HD suggesting that STEP inhibition could be a good therapeutic strategy in HD patients.
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Cognición/fisiología , Modelos Animales de Enfermedad , Enfermedad de Huntington/metabolismo , Destreza Motora/fisiología , Farmacogenética/métodos , Proteínas Tirosina Fosfatasas no Receptoras/deficiencia , Animales , Enfermedad de Huntington/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Actividad Motora/fisiología , Farmacogenética/tendencias , Proteínas Tirosina Fosfatasas no Receptoras/genéticaRESUMEN
The progressive restriction of differentiation potential from pluripotent embryonic stem cells (ESCs) to tissue-specific stem cells involves widespread epigenetic reprogramming, including modulation of DNA methylation patterns. Skeletal muscle stem cells are required for the growth, maintenance, and regeneration of skeletal muscle. To investigate the contribution of DNA methylation to the establishment of the myogenic program, we analyzed ESCs, skeletal muscle stem cells in proliferating (myoblasts) and differentiating conditions (myotubes), and mature myofibers. About 1.000 differentially methylated regions were identified during muscle-lineage determination and terminal differentiation, mainly located in gene bodies and intergenic regions. As a whole, myogenic stem cells showed a gain of DNA methylation, while muscle differentiation was accompanied by loss of DNA methylation in CpG-poor regions. Notably, the hypomethylated regions in myogenic stem cells were neighbored by enhancer-type chromatin, suggesting the involvement of DNA methylation in the regulation of cell-type specific enhancers. Interestingly, we demonstrated the hypomethylation of the muscle cell-identity Myf5 super-enhancer only in muscle cells. Furthermore, we observed that upstream stimulatory factor 1 binding to Myf5 super-enhancer occurs upon DNA demethylation in myogenic stem cells. Taken altogether, we characterized the unique DNA methylation signature of skeletal muscle stem cells and highlighted the importance of DNA methylation-mediated regulation of cell identity Myf5 super-enhancer during cellular differentiation.
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Diferenciación Celular/genética , Linaje de la Célula/genética , Metilación de ADN/genética , Desarrollo de Músculos/genética , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/citología , Regulación de la Expresión Génica/genética , Células Madre Embrionarias Humanas/metabolismo , Humanos , Proteínas Musculares/genéticaRESUMEN
Alterations in motor development often accompany neurodevelopmental disorders (NDD) and can have an impact on social interaction and communication. Studying motor development and function in mouse models of NDDs can offer a window to identify underlying biological mechanisms and establish preclinical outcome measures for testing therapeutics. This chapter describes tests to measure motor developmental milestones early postnatally and adult motor functions in mouse models of NDDs.
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Modelos Animales de Enfermedad , Trastornos del Neurodesarrollo , Animales , Trastornos del Neurodesarrollo/fisiopatología , Trastornos del Neurodesarrollo/patología , Ratones , Actividad Motora , HumanosRESUMEN
Monozygotic (MZ) twins are partially concordant for most complex diseases, including autoimmune disorders. Whereas phenotypic concordance can be used to study heritability, discordance suggests the role of non-genetic factors. In autoimmune diseases, environmentally driven epigenetic changes are thought to contribute to their etiology. Here we report the first high-throughput and candidate sequence analyses of DNA methylation to investigate discordance for autoimmune disease in twins. We used a cohort of MZ twins discordant for three diseases whose clinical signs often overlap: systemic lupus erythematosus (SLE), rheumatoid arthritis, and dermatomyositis. Only MZ twins discordant for SLE featured widespread changes in the DNA methylation status of a significant number of genes. Gene ontology analysis revealed enrichment in categories associated with immune function. Individual analysis confirmed the existence of DNA methylation and expression changes in genes relevant to SLE pathogenesis. These changes occurred in parallel with a global decrease in the 5-methylcytosine content that was concomitantly accompanied with changes in DNA methylation and expression levels of ribosomal RNA genes, although no changes in repetitive sequences were found. Our findings not only identify potentially relevant DNA methylation markers for the clinical characterization of SLE patients but also support the notion that epigenetic changes may be critical in the clinical manifestations of autoimmune disease.
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Metilación de ADN , Enfermedades en Gemelos/genética , Lupus Eritematoso Sistémico/genética , Gemelos Monocigóticos/genética , 5-Metilcitosina/química , Artritis Reumatoide/genética , Enfermedades Autoinmunes/genética , Estudios de Cohortes , Islas de CpG/genética , Dermatomiositis/genética , Femenino , Genes de ARNr , Humanos , Masculino , Regiones Promotoras Genéticas , Análisis de Secuencia de ADN/métodosRESUMEN
Abnormalities in neocortical and synaptic development are linked to neurodevelopmental disorders. However, the molecular and cellular mechanisms governing initial synapse formation in the prenatal neocortex remain poorly understood. Using polysome profiling coupled with snRNAseq on human cortical samples at various fetal phases, we identify human mRNAs, including those encoding synaptic proteins, with finely controlled translation in distinct cell populations of developing frontal neocortices. Examination of murine and human neocortex reveals that the RNA binding protein and translational regulator, CELF4, is expressed in compartments enriched in initial synaptogenesis: the marginal zone and the subplate. We also find that Celf4/CELF4-target mRNAs are encoded by risk genes for adverse neurodevelopmental outcomes translating into synaptic proteins. Surprisingly, deleting Celf4 in the forebrain disrupts the balance of subplate synapses in a sex-specific fashion. This highlights the significance of RNA binding proteins and mRNA translation in evolutionarily advanced synaptic development, potentially contributing to sex differences.
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Proteínas CELF , Neocórtex , Animales , Femenino , Humanos , Masculino , Ratones , Embarazo , Neocórtex/metabolismo , Neuronas/metabolismo , Polirribosomas/metabolismo , Biosíntesis de Proteínas , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Sinapsis/metabolismo , Proteínas CELF/genética , Proteínas CELF/metabolismoRESUMEN
We conducted a large-scale study of whole-brain morphometry, analyzing 3.7 peta-voxels of mouse brain images at the single-cell resolution, producing one of the largest multi-morphometry databases of mammalian brains to date. We spatially registered 205 mouse brains and associated data from six Brain Initiative Cell Census Network (BICCN) data sources covering three major imaging modalities from five collaborative projects to the Allen Common Coordinate Framework (CCF) atlas, annotated 3D locations of cell bodies of 227,581 neurons, modeled 15,441 dendritic microenvironments, characterized the full morphology of 1,891 neurons along with their axonal motifs, and detected 2.58 million putative synaptic boutons. Our analysis covers six levels of information related to neuronal populations, dendritic microenvironments, single-cell full morphology, sub-neuronal dendritic and axonal arborization, axonal boutons, and structural motifs, along with a quantitative characterization of the diversity and stereotypy of patterns at each level. We identified 16 modules consisting of highly intercorrelated brain regions in 13 functional brain areas corresponding to 314 anatomical regions in CCF. Our analysis revealed the dendritic microenvironment as a powerful method for delineating brain regions of cell types and potential subtypes. We also found that full neuronal morphologies can be categorized into four distinct classes based on spatially tuned morphological features, with substantial cross-areal diversity in apical dendrites, basal dendrites, and axonal arbors, along with quantified stereotypy within cortical, thalamic and striatal regions. The lamination of somas was found to be more effective in differentiating neuron arbors within the cortex. Further analysis of diverging and converging projections of individual neurons in 25 regions throughout the brain reveals branching preferences in the brain-wide and local distributions of axonal boutons. Overall, our study provides a comprehensive description of key anatomical structures of neurons and their types, covering a wide range of scales and features, and contributes to our understanding of neuronal diversity and its function in the mammalian brain.
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RTP801/REDD1 is a stress-regulated protein whose levels are increased in several neurodegenerative diseases such as Parkinson's, Alzheimer's, and Huntington's diseases (HD). RTP801 downregulation ameliorates behavioral abnormalities in several mouse models of these disorders. In HD, RTP801 mediates mutant huntingtin (mhtt) toxicity in in vitro models and its levels are increased in human iPSCs, human postmortem putamen samples, and in striatal synaptosomes from mouse models of the disease. Here, we investigated the role of RTP801 in the hippocampal pathophysiology of HD. We found that RTP801 levels are increased in the hippocampus of HD patients in correlation with gliosis markers. Although RTP801 expression is not altered in the hippocampus of the R6/1 mouse model of HD, neuronal RTP801 silencing in the dorsal hippocampus with shRNA containing AAV particles ameliorates cognitive alterations. This recovery is associated with a partial rescue of synaptic markers and with a reduction in inflammatory events, especially microgliosis. Altogether, our results indicate that RTP801 could be a marker of hippocampal neuroinflammation in HD patients and a promising therapeutic target of the disease.
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Disfunción Cognitiva , Enfermedad de Huntington , Animales , Modelos Animales de Enfermedad , Humanos , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Ratones , Ratones Transgénicos , Enfermedades NeuroinflamatoriasRESUMEN
BACKGROUND: Mutations in the X-linked gene DDX3X account for approximately 2% of intellectual disability in females, often comorbid with behavioral problems, motor deficits, and brain malformations. DDX3X encodes an RNA helicase with emerging functions in corticogenesis and synaptogenesis. METHODS: We generated a Ddx3x haploinsufficient mouse (Ddx3x+/- females) with construct validity for DDX3X loss-of-function mutations. We used standardized batteries to assess developmental milestones and adult behaviors, as well as magnetic resonance imaging and immunostaining of cortical projection neurons to capture early postnatal changes in brain development. RESULTS: Ddx3x+/- females showed physical, sensory, and motor delays that evolved into behavioral anomalies in adulthood, including hyperactivity, anxiety-like behaviors, cognitive impairments in specific tasks (e.g., contextual fear memory but not novel object recognition memory), and motor deficits. Motor function declined with age but not if mice were previously exposed to behavioral training. Developmental and behavioral changes were associated with a reduction in brain volume, with some regions (e.g., cortex and amygdala) disproportionally affected. Cortical thinning was accompanied by defective cortical lamination, indicating that Ddx3x regulates the balance of glutamatergic neurons in the developing cortex. CONCLUSIONS: These data shed new light on the developmental mechanisms driving DDX3X syndrome and support construct and face validity of this novel preclinical mouse model.
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Discapacidad Intelectual , Animales , ARN Helicasas DEAD-box/genética , Modelos Animales de Enfermedad , Femenino , Ratones , Neurogénesis , Fenotipo , SíndromeRESUMEN
Lamins are crucial proteins for nuclear functionality. Here, we provide new evidence showing that increased lamin B1 levels contribute to the pathophysiology of Huntington's disease (HD), a CAG repeat-associated neurodegenerative disorder. Through fluorescence-activated nuclear suspension imaging, we show that nucleus from striatal medium-sized spiny and CA1 hippocampal neurons display increased lamin B1 levels, in correlation with altered nuclear morphology and nucleocytoplasmic transport disruption. Moreover, ChIP-sequencing analysis shows an alteration of lamin-associated chromatin domains in hippocampal nuclei, accompanied by changes in chromatin accessibility and transcriptional dysregulation. Supporting lamin B1 alterations as a causal role in mutant huntingtin-mediated neurodegeneration, pharmacological normalization of lamin B1 levels in the hippocampus of the R6/1 mouse model of HD by betulinic acid administration restored nuclear homeostasis and prevented motor and cognitive dysfunction. Collectively, our work points increased lamin B1 levels as a new pathogenic mechanism in HD and provides a novel target for its intervention.
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Enfermedad de Huntington , Animales , Cuerpo Estriado , Enfermedad de Huntington/genética , Lamina Tipo B/genética , Ratones , NeuronasRESUMEN
Autism spectrum disorder (ASD) is a prevalent neurodevelopmental disorder characterized by impairments in social communication and social interaction, and the presence of repetitive behaviors and/or restricted interests. In the past few years, large-scale whole-exome sequencing and genome-wide association studies have made enormous progress in our understanding of the genetic risk architecture of ASD. While showing a complex and heterogeneous landscape, these studies have led to the identification of genetic loci associated with ASD risk. The intersection of genetic and transcriptomic analyses have also begun to shed light on functional convergences between risk genes, with the mid-fetal development of the cerebral cortex emerging as a critical nexus for ASD. In this review, we provide a concise summary of the latest genetic discoveries on ASD. We then discuss the studies in postmortem tissues, stem cell models, and rodent models that implicate recently identified ASD risk genes in cortical development.
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Trastorno Autístico/genética , Malformaciones del Desarrollo Cortical/genética , Animales , Modelos Animales de Enfermedad , Humanos , Ratones , Factores de RiesgoRESUMEN
Mouse models of intestinal crypt cell differentiation and tumorigenesis have been used to characterize the molecular mechanisms underlying both processes. DNA methylation is a key epigenetic mark and plays an important role in cell identity and differentiation programs and cancer. To get insights into the dynamics of cell differentiation and malignant transformation we have compared the DNA methylation profiles along the mouse small intestine crypt and early stages of tumorigenesis. Genome-scale analysis of DNA methylation together with microarray gene expression have been applied to compare intestinal crypt stem cells (EphB2high), differentiated cells (EphB2negative), ApcMin/+ adenomas and the corresponding non-tumor adjacent tissue, together with small and large intestine samples and the colon cancer cell line CT26. Compared with late stages, small intestine crypt differentiation and early stages of tumorigenesis display few and relatively small changes in DNA methylation. Hypermethylated loci are largely shared by the two processes and affect the proximities of promoter and enhancer regions, with enrichment in genes associated with the intestinal stem cell signature and the PRC2 complex. The hypermethylation is progressive, with minute levels in differentiated cells, as compared with intestinal stem cells, and reaching full methylation in advanced stages. Hypomethylation shows different signatures in differentiation and cancer and is already present in the non-tumor tissue adjacent to the adenomas in ApcMin/+ mice, but at lower levels than advanced cancers. This study provides a reference framework to decipher the mechanisms driving mouse intestinal tumorigenesis and also the human counterpart.