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1.
Sci Adv ; 10(5): eadk3060, 2024 Feb 02.
Artículo en Inglés | MEDLINE | ID: mdl-38306432

RESUMEN

Effective, unbiased, high-throughput methods to functionally identify both class II and class I HLA-presented T cell epitopes and their cognate T cell receptors (TCRs) are essential for and prerequisite to diagnostic and therapeutic applications, yet remain underdeveloped. Here, we present T-FINDER [T cell Functional Identification and (Neo)-antigen Discovery of Epitopes and Receptors], a system to rapidly deconvolute CD4 and CD8 TCRs and targets physiologically processed and presented by an individual's unmanipulated, complete human leukocyte antigen (HLA) haplotype. Combining a highly sensitive TCR signaling reporter with an antigen processing system to overcome previously undescribed limitations to target expression, T-FINDER both robustly identifies unknown peptide:HLA ligands from antigen libraries and rapidly screens and functionally validates the specificity of large TCR libraries against known or predicted targets. To demonstrate its capabilities, we apply the platform to multiple TCR-based applications, including diffuse midline glioma, celiac disease, and rheumatoid arthritis, providing unique biological insights and showcasing T-FINDER's potency and versatility.


Asunto(s)
Antígenos de Histocompatibilidad Clase I , Receptores de Antígenos de Linfocitos T , Humanos , Ligandos , Receptores de Antígenos de Linfocitos T/metabolismo , Antígenos HLA , Antígenos de Histocompatibilidad Clase II
2.
Arthritis Rheumatol ; 75(5): 673-684, 2023 05.
Artículo en Inglés | MEDLINE | ID: mdl-36409582

RESUMEN

OBJECTIVE: CD4+ T cells are implicated in rheumatoid arthritis (RA) pathology from the strong association between RA and certain HLA class II gene variants. This study was undertaken to examine the synovial T cell receptor (TCR) repertoire, T cell phenotypes, and T cell specificities in small joints of RA patients at time of diagnosis before therapeutic intervention. METHODS: Sixteen patients, of whom 11 patients were anti-citrullinated protein antibody (ACPA)-positive and 5 patients were ACPA-, underwent ultrasound-guided synovial biopsy of a small joint (n = 13) or arthroscopic synovial biopsy of a large joint (n = 3), followed by direct sorting of single T cells for paired sequencing of the αß TCR together with flow cytometry analysis. TCRs from expanded CD4+ T cell clones of 4 patients carrying an HLA-DRB1*04:01 allele were artificially reexpressed to study antigen specificity. RESULTS: T cell analysis demonstrated CD4+ dominance and the presence of peripheral helper T-like cells in both patient groups. We identified >4,000 unique TCR sequences, as well as 225 clonal expansions. Additionally, T cells with double α-chains were a recurring feature. We identified a biased gene usage of the Vß chain segment TRBV20-1 in CD4+ cells from ACPA+ patients. In vitro stimulation of T cell lines expressing selected TCRs with an extensive panel of citrullinated and viral peptides identified several different virus-specific TCRs (e.g., human cytomegalovirus and human herpesvirus 2). Still, the majority of clones remained orphans with unknown specificity. CONCLUSION: Minimally invasive biopsies of the RA synovium allow for single-cell TCR sequencing and phenotyping. Clonally expanded, viral-reactive T cells account for part of the diverse CD4+ T cell repertoire. TRBV20-1 bias in ACPA+ patients suggests recognition of common antigens.


Asunto(s)
Artritis Reumatoide , Humanos , Membrana Sinovial/patología , Linfocitos T CD4-Positivos , Receptores de Antígenos de Linfocitos T/genética , Cadenas HLA-DRB1/genética
3.
Genome Biol ; 23(1): 55, 2022 02 16.
Artículo en Inglés | MEDLINE | ID: mdl-35172874

RESUMEN

BACKGROUND: Multiplexing of samples in single-cell RNA-seq studies allows a significant reduction of the experimental costs, straightforward identification of doublets, increased cell throughput, and reduction of sample-specific batch effects. Recently published multiplexing techniques using oligo-conjugated antibodies or -lipids allow barcoding sample-specific cells, a process called "hashing." RESULTS: Here, we compare the hashing performance of TotalSeq-A and -C antibodies, custom synthesized lipids and MULTI-seq lipid hashes in four cell lines, both for single-cell RNA-seq and single-nucleus RNA-seq. We also compare TotalSeq-B antibodies with CellPlex reagents (10x Genomics) on human PBMCs and TotalSeq-B with different lipids on primary mouse tissues. Hashing efficiency was evaluated using the intrinsic genetic variation of the cell lines and mouse strains. Antibody hashing was further evaluated on clinical samples using PBMCs from healthy and SARS-CoV-2 infected patients, where we demonstrate a more affordable approach for large single-cell sequencing clinical studies, while simultaneously reducing batch effects. CONCLUSIONS: Benchmarking of different hashing strategies and computational pipelines indicates that correct demultiplexing can be achieved with both lipid- and antibody-hashed human cells and nuclei, with MULTISeqDemux as the preferred demultiplexing function and antibody-based hashing as the most efficient protocol on cells. On nuclei datasets, lipid hashing delivers the best results. Lipid hashing also outperforms antibodies on cells isolated from mouse brain. However, antibodies demonstrate better results on tissues like spleen or lung.


Asunto(s)
COVID-19/sangre , Análisis de Secuencia de ARN/métodos , Análisis de la Célula Individual/métodos , Animales , Anticuerpos/química , Estudios de Casos y Controles , Línea Celular Tumoral , Núcleo Celular/química , Humanos , Lípidos/química , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Neutrófilos/química , Neutrófilos/inmunología , Neutrófilos/virología
4.
Allergy ; 76(6): 1825-1835, 2021 06.
Artículo en Inglés | MEDLINE | ID: mdl-33150583

RESUMEN

BACKGROUND: Atabecestat is an orally administered BACE inhibitor developed to treat Alzheimer's disease. Elevations in hepatic enzymes were detected in a number of in trial patients, which resulted in termination of the drug development programme. Immunohistochemical characterization of liver tissue from an index case of atabecestat-mediated liver injury revealed an infiltration of T-lymphocytes in areas of hepatocellular damage. This coupled with the fact that liver injury had a delayed onset suggests that the adaptive immune system may be involved in the pathogenesis. The aim of this study was to generate and characterize atabecestat(metabolite)-responsive T-cell clones from patients with liver injury. METHODS: Peripheral blood mononuclear cells were cultured with atabecestat and its metabolites (diaminothiazine [DIAT], N-acetyl DIAT & epoxide) and cloning was attempted in a number of patients. Atabecestat(metabolite)-responsive clones were analysed in terms of T-cell phenotype, function, pathways of T-cell activation and cross-reactivity with structurally related compounds. RESULTS: CD4+ T-cell clones activated with the DIAT metabolite were detected in 5 out of 8 patients (up to 4.5% cloning efficiency). Lower numbers of CD4+ and CD8+ clones displayed reactivity against atabecestat. Clones proliferated and secreted IFN-γ, IL-13 and cytolytic molecules following atabecestat or DIAT stimulation. Certain atabecestat and DIAT-responsive clones cross-reacted with N-acetyl DIAT; however, no cross-reactivity was observed between atabecestat and DIAT. CD4+ clones were activated through a direct, reversible compound-HLA class II interaction with no requirement for protein processing. CONCLUSION: The detection of atabecestat metabolite-responsive T-cell clones activated via a pharmacological interactions pathway in patients with liver injury is indicative of an immune-based mechanism for the observed hepatic enzyme elevations.


Asunto(s)
Preparaciones Farmacéuticas , Linfocitos T , Linfocitos T CD4-Positivos , Células Clonales , Humanos , Leucocitos Mononucleares , Hígado , Activación de Linfocitos , Piridinas , Tiazinas
5.
Methods Mol Biol ; 1806: 233-253, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-29956280

RESUMEN

Currently one of the few molecules that equally excites a neuroscientist, a cancer biologist, an immunologist, and a developmental biologist is progranulin (GRN/Grn)-a pluripotent growth factor that plays key roles in cell survival, proliferation, development, tissue regeneration, inflammation, wound healing, and angiogenesis. However, the molecular pathways associated with GRN signaling involved in these varied physiological processes are not understood. Gene inactivation has been considered as one of the best methods to delineate the biological role of a protein, and gene targeting is a direct means to disrupt a gene's open reading frame and block its expression, for instance, in a mouse. Such a gene knockout animal model also served as an in vivo disease model where loss of gene or its function is thought to be the primary disease mechanism, as is the case with progranulin loss of function in frontotemporal lobar degeneration (FTLD). It is estimated that up to half of the cases of familial, dominant FTLD might be due to GRN haploinsufficiency. To understand the molecular pathways associated with GRN loss, constitutive and conditional progranulin knockout (Grn-/-) mice have also been constructed in several laboratories, including ours. These mice show several disease-characteristic features and suggest that continued studies on the Grn-/- mice would be instructive in the understanding of complex GRN biology in health and disease.


Asunto(s)
Conducta Animal , Encéfalo/metabolismo , Biología Molecular/métodos , Progranulinas/metabolismo , Envejecimiento , Animales , Aprendizaje por Laberinto , Ratones Noqueados , Proteínas del Tejido Nervioso/metabolismo , Fosforilación , Reacción en Cadena de la Polimerasa , Progranulinas/deficiencia , Progranulinas/aislamiento & purificación
6.
Mol Neurobiol ; 48(1): 22-35, 2013 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-23475610

RESUMEN

Mutations in TAR DNA-binding protein 43 (TDP-43) are associated with familial forms of amyotrophic lateral sclerosis (ALS), while wild-type TDP-43 is a pathological hallmark of patients with sporadic ALS and frontotemporal lobar degeneration (FTLD). Various in vitro and in vivo studies have also demonstrated toxicity of both mutant and wild-type TDP-43 to neuronal cells. To study the potential additional toxicity incurred by mutant TDP-43 in vivo, we generated mutant human TDP-43 (p.M337V) transgenic mouse lines driven by the Thy-1.2 promoter (Mt-TAR) and compared them in the same experimental setting to the disease phenotype observed in wild-type TDP-43 transgenic lines (Wt-TAR) expressing comparable TDP-43 levels. Overexpression of mutant TDP-43 leads to a worsened dose-dependent disease phenotype in terms of motor dysfunction, neurodegeneration, gliosis, and development of ubiquitin and phosphorylated TDP-43 pathology. Furthermore, we show that cellular aggregate formation or accumulation of TDP-43 C-terminal fragments (CTFs) are not primarily responsible for development of the observed disease phenotype in both mutant and wild-type TDP-43 mice.


Asunto(s)
Sustitución de Aminoácidos/genética , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/patología , Proteínas de Unión al ADN/genética , Progresión de la Enfermedad , Animales , Núcleo Celular/metabolismo , Núcleo Celular/ultraestructura , Eosinófilos/metabolismo , Eosinófilos/ultraestructura , Gliosis/patología , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Mitocondrias/metabolismo , Mitocondrias/ultraestructura , Proteínas Mutantes/metabolismo , Neuronas/patología , Neuronas/ultraestructura , Fenotipo , Fosforilación , Procesamiento Proteico-Postraduccional , Médula Espinal/patología , Médula Espinal/ultraestructura , Transgenes/genética , Proteínas Ubiquitinadas/metabolismo
7.
J Pathol ; 228(1): 67-76, 2012 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-22733568

RESUMEN

Loss-of-function mutations in progranulin (GRN) are associated with frontotemporal lobar degeneration with intraneuronal ubiquitinated protein accumulations composed primarily of hyperphosphorylated TDP-43 (FTLD-TDP). The mechanism by which GRN deficiency causes TDP-43 pathology or neurodegeneration remains elusive. To explore the role of GRN in vivo, we established Grn knockout mice using a targeted genomic recombination approach and Cre-LoxP technology. Constitutive Grn homozygous knockout (Grn(-/-) ) mice were born in an expected Mendelian pattern of inheritance and showed no phenotypic alterations compared to heterozygous (Grn(+/-) ) or wild-type (Wt) littermates until 10 months of age. From then, Grn(-/-) mice showed reduced survival accompanied by significantly increased gliosis and ubiquitin-positive accumulations in the cortex, hippocampus, and subcortical regions. Although phosphorylated TDP-43 could not be detected in the ubiquitinated inclusions, elevated levels of hyperphosphorylated full-length TDP-43 were recovered from detergent-insoluble brain fractions of Grn(-/-) mice. Phosphorylated TDP-43 increased with age and was primarily extracted from the nuclear fraction. Grn(-/-) mice also showed degenerative liver changes and cathepsin D-positive foamy histiocytes within sinusoids, suggesting widespread defects in lysosomal turnover. An increase in insulin-like growth factor (IGF)-1 was observed in Grn(-/-) brains, and increased IGF-1 signalling has been associated with decreased longevity. Our data suggest that progranulin deficiency in mice leads to reduced survival in adulthood and increased cellular ageing accompanied by hyperphosphorylation of TDP-43, and recapitulates key aspects of FTLD-TDP neuropathology.


Asunto(s)
Senescencia Celular , Proteínas de Unión al ADN/metabolismo , Degeneración Lobar Frontotemporal/patología , Péptidos y Proteínas de Señalización Intercelular/fisiología , Animales , Conducta Animal/fisiología , Encéfalo/metabolismo , Encéfalo/patología , Proteínas de Unión al ADN/genética , Femenino , Degeneración Lobar Frontotemporal/genética , Degeneración Lobar Frontotemporal/mortalidad , Expresión Génica , Gliosis/metabolismo , Gliosis/patología , Granulinas , Hígado/patología , Longevidad/fisiología , Masculino , Ratones , Ratones Noqueados , Fosforilación , Progranulinas , Tasa de Supervivencia , Ubiquitina/metabolismo
8.
Biochem Soc Trans ; 39(4): 954-9, 2011 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-21787329

RESUMEN

TDP-43 (TAR DNA-binding protein 43) has been identified as a key protein of ubiquitinated inclusions in brains of patients with ALS (amyotrophic lateral sclerosis) or FTLD (frontotemporal lobar degeneration), defining a new pathological disease spectrum. Recently, coding mutations have been identified in the TDP-43 gene (TARDBP), which further confirmed the pathogenic nature of the protein. Today, several animal models have been generated to gain more insight into the disease-causing pathways of the FTLD/ALS spectrum. This mini-review summarizes the current status of TDP-43 models, with a focus on mutant TDP-43.


Asunto(s)
Esclerosis Amiotrófica Lateral/genética , Proteínas de Unión al ADN/genética , Modelos Animales de Enfermedad , Degeneración Lobar Frontotemporal/genética , Proteínas Mutantes/genética , Sustitución de Aminoácidos , Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/patología , Animales , Proteínas de Unión al ADN/metabolismo , Degeneración Lobar Frontotemporal/metabolismo , Humanos , Proteínas Mutantes/metabolismo
9.
J Neurochem ; 115(3): 735-47, 2010 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-20731760

RESUMEN

Null mutations in progranulin (GRN) are associated with frontotemporal lobar degeneration characterized by intraneuronal accumulation of TAR DNA-binding protein-43 (TDP-43). However, the mechanism by which GRN deficiency leads to neurodegeneration remains largely unknown. In primary cortical neurons derived from Grn knockout (Grn(-/-) ) mice, we found that Grn-deficiency causes significantly reduced neuronal survival and increased caspase-mediated apoptosis, which was not observed in primary mouse embryonic fibroblasts derived from Grn(-/-) mice. Also, neurons derived from Grn(-/-) mice showed an increased amount of pTDP-43 accumulations. Furthermore, proteasomal inhibition with MG132 caused increased caspase-mediated TDP-43 fragmentation and accumulation of detergent-insoluble 35- and 25-kDa C-terminal fragments in Grn(-/-) neurons and mouse embryonic fibroblasts. Interestingly, full-length TDP-43 also accumulated in the detergent-insoluble fraction, and caspase-inhibition prevented MG132-induced generation of TDP-43 C-terminal fragments but did not block the pathological conversion of full-length TDP-43 from soluble to insoluble species. These data suggest that GRN functions as a survival factor for cortical neurons and GRN-deficiency causes increased susceptibility to cellular stress. This leads to increased aggregation and accumulation of full-length TDP-43 along with its C-terminal derivatives by both caspase-dependent and independent mechanisms.


Asunto(s)
Caspasas/metabolismo , Corteza Cerebral/metabolismo , Proteínas de Unión al ADN/metabolismo , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Animales , Apoptosis/efectos de los fármacos , Western Blotting , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Corteza Cerebral/citología , Corteza Cerebral/efectos de los fármacos , Proteínas de Unión al ADN/química , Activación Enzimática/fisiología , Fibroblastos/metabolismo , Granulinas , Inmunohistoquímica , Ratones , Ratones Noqueados , Proteína Oncogénica v-akt/metabolismo , Fosforilación , Progranulinas , Complejo de la Endopetidasa Proteasomal , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/farmacología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
10.
Proc Natl Acad Sci U S A ; 107(8): 3858-63, 2010 Feb 23.
Artículo en Inglés | MEDLINE | ID: mdl-20133711

RESUMEN

Neuronal cytoplasmic and intranuclear aggregates of RNA-binding protein TDP-43 are a hallmark feature of neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). ALS and FTLD show a considerable clinical and pathological overlap and occur as both familial and sporadic forms. Though missense mutations in TDP-43 cause rare forms of familial ALS, it is not yet known whether this is due to loss of TDP-43 function or gain of aberrant function. Moreover, the role of wild-type (WT) TDP-43, associated with the majority of familial and sporadic ALS/FTLD patients, is also currently unknown. Generating homozygous and hemizygous WT human TDP-43 transgenic mouse lines, we show here a dose-dependent degeneration of cortical and spinal motor neurons and development of spastic quadriplegia reminiscent of ALS. A dose-dependent degeneration of nonmotor cortical and subcortical neurons characteristic of FTLD was also observed. Neurons in the affected spinal cord and brain regions showed accumulation of TDP-43 nuclear and cytoplasmic aggregates that were both ubiquitinated and phosphorylated as observed in ALS/FTLD patients. Moreover, the characteristic approximately 25-kDa C-terminal fragments (CTFs) were also recovered from nuclear fractions and correlated with disease development and progression in WT TDP-43 mice. These findings suggest that approximately 25-kDa TDP-43 CTFs are noxious to neurons by a gain of aberrant nuclear function.


Asunto(s)
Esclerosis Amiotrófica Lateral/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Degeneración Lobar Frontotemporal/genética , Cuerpos de Inclusión/metabolismo , Parálisis/genética , Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/patología , Animales , Degeneración Lobar Frontotemporal/metabolismo , Degeneración Lobar Frontotemporal/patología , Humanos , Cuerpos de Inclusión/genética , Ratones , Ratones Transgénicos , Espasticidad Muscular/genética , Espasticidad Muscular/patología , Mutación Missense , Parálisis/patología
11.
J Pathol ; 219(2): 173-81, 2009 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-19557827

RESUMEN

Amyloid-beta (Abeta) plaques are pathological hallmarks of Alzheimer disease (AD). In addition, innate inflammatory responses, such as those mediated by microglia, are integral to the pathogenesis of AD. Interestingly, only dense-core plaques and not diffuse plaques are associated with neuritic and inflammatory pathology in AD patients as well as in mouse AD models. However, the precise neuropathological changes that occur in the brain in response to amyloid deposition are largely unknown. To study the molecular mechanism(s) responsible for Abeta-mediated neuropathology, we performed a gene expression analysis on laser-microdissected brain tissue of Tg2576 and APPPS1 mice that are characterized by different types of amyloid plaques and genetic backgrounds. Data were validated by image and biochemical analyses on different ages of Tg2576, APPPS1, and Abeta42-depositing BRI-Abeta42 mice. Consistent with an important role of inflammatory responses in AD, we identified progranulin (mouse Grn; human GRN) as one of the top ten up-regulated molecules in Tg2576 ( approximately 8-fold increased) and APPPS1 ( approximately 2-fold increased) mice compared to littermate controls, and among the eight significantly up-regulated molecules common to both mouse models. In addition, Grn levels correlated significantly with amyloid load, especially the dense-core plaque pathology (p < 0.001). We further showed that Grn is up-regulated in microglia and neurons and neurites around dense-core plaques, but not in astrocytes or oligodendrocytes, as has been shown in AD patients. Our data therefore support the ongoing use of these mouse models in drug trials, especially those with anti-inflammatory compounds. Moreover, the correlation of Grn with increasing disease severity in AD mouse models prompts human studies exploring the viability of GRN as a disease biomarker. Because loss of GRN has recently been shown to cause frontotemporal dementia and serves as a risk factor for AD, the strong GRN reactivity around dense-core plaques is consistent with an important role of this factor in AD pathogenesis.


Asunto(s)
Enfermedad de Alzheimer/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Placa Amiloide/metabolismo , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/metabolismo , Animales , Encéfalo/metabolismo , Encéfalo/patología , Modelos Animales de Enfermedad , Femenino , Perfilación de la Expresión Génica/métodos , Granulinas , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microglía/metabolismo , Neuritas/metabolismo , Neuronas/metabolismo , Placa Amiloide/patología , Progranulinas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Regulación hacia Arriba
12.
Neurobiol Aging ; 29(2): 241-52, 2008 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-17112635

RESUMEN

Transgenic mouse models of Alzheimer's disease (AD) expressing high levels of amyloid precursor protein (APP) with familial AD (FAD) mutations have proven to be extremely useful in understanding pathogenic processes of AD especially those that involve amyloidogenesis. We earlier described Austrian APP T714I pathology that leads to one of the earliest AD age-at-onsets with abundant intracellular and extracellular amyloid deposits in brain. The latter strikingly was non-fibrillar diffuse amyloid, composed of N-truncated A beta 42 in absence of A beta 40. In vitro, this mutation leads to one of the highest A beta 42/A beta 40 ratios among all FAD mutations. We generated an APP T714I transgenic mouse model that despite having 10 times lower transgene than endogenous murine APP deposited intraneuronal A beta in brain by 6 months of age. Accumulations increased with age, and this was paralleled by decreased brain sizes on volumetric MRI, compared to age-matched and similar transgene-expressing APP wild-type mice, although, with these levels of transgenic expression we did not detect neuronal loss or significant memory impairment. Immunohistochemical studies revealed that the majority of the intraneuronal A beta deposits colocalized with late endosomal markers, although some A beta inclusions were also positive for lysosomal and Golgi markers. These data support earlier observations of A beta accumulation in the endosomal-lysosomal pathway and the hypothesis that intraneuronal accumulation of A beta could be an important factor in the AD pathogenesis.


Asunto(s)
Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/genética , Encéfalo/patología , Isoleucina/genética , Tirosina/genética , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/fisiopatología , Análisis de Varianza , Animales , Conducta Animal/fisiología , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Humanos , Imagen por Resonancia Magnética , Aprendizaje por Laberinto/fisiología , Ratones , Ratones Transgénicos , Neuronas/citología , Fragmentos de Péptidos/metabolismo , Desempeño Psicomotor/fisiología
13.
Nature ; 442(7105): 920-4, 2006 Aug 24.
Artículo en Inglés | MEDLINE | ID: mdl-16862115

RESUMEN

Frontotemporal dementia (FTD) with ubiquitin-immunoreactive neuronal inclusions (both cytoplasmic and nuclear) of unknown nature has been linked to a chromosome 17q21 region (FTDU-17) containing MAPT (microtubule-associated protein tau). FTDU-17 patients have consistently been shown to lack a tau-immunoreactive pathology, a feature characteristic of FTD with parkinsonism linked to mutations in MAPT (FTDP-17). Furthermore, in FTDU-17 patients, mutations in MAPT and genomic rearrangements in the MAPT region have been excluded by both genomic sequencing and fluorescence in situ hybridization on mechanically stretched chromosomes. Here we demonstrate that FTDU-17 is caused by mutations in the gene coding for progranulin (PGRN), a growth factor involved in multiple physiological and pathological processes including tumorigenesis. Besides the production of truncated PGRN proteins due to premature stop codons, we identified a mutation within the splice donor site of intron 0 (IVS0 + 5G > C), indicating loss of the mutant transcript by nuclear degradation. The finding was made within an extensively documented Belgian FTDU-17 founder family. Transcript and protein analyses confirmed the absence of the mutant allele and a reduction in the expression of PGRN. We also identified a mutation (c.3G > A) in the Met1 translation initiation codon, indicating loss of PGRN due to lack of translation of the mutant allele. Our data provide evidence that PGRN haploinsufficiency leads to neurodegeneration because of reduced PGRN-mediated neuronal survival. Furthermore, in a Belgian series of familial FTD patients, PGRN mutations were 3.5 times more frequent than mutations in MAPT, underscoring a principal involvement of PGRN in FTD pathogenesis.


Asunto(s)
Cromosomas Humanos Par 17/genética , Demencia/genética , Lóbulo Frontal/fisiopatología , Péptidos y Proteínas de Señalización Intercelular/deficiencia , Mutación/genética , Lóbulo Temporal/fisiopatología , Ubiquitina/metabolismo , Bélgica , Análisis Mutacional de ADN , Demencia/fisiopatología , Lóbulo Frontal/metabolismo , Ligamiento Genético/genética , Humanos , Péptidos y Proteínas de Señalización Intercelular/genética , Mapeo Físico de Cromosoma , Progranulinas , Sitios de Empalme de ARN/genética , Lóbulo Temporal/metabolismo
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