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1.
Biol Psychiatry ; 2024 Jul 17.
Artículo en Inglés | MEDLINE | ID: mdl-39029776

RESUMEN

BACKGROUND: Parkinson's disease (PD) is the second most common neurodegenerative disease, following Alzheimer's. It is characterized by the aggregation of α-synuclein into Lewy bodies and Lewy neurites in the brain. Microglia-driven neuroinflammation may contribute to neuronal death in PD, however the exact role of microglia remains unclear and has been understudied. The A53T mutation in the gene coding for α-synuclein has been linked to early-onset PD, and exposure to A53T-mutant human α-synuclein increases the potential for inflammation of murine microglia. To date, its effect has not been studied in human microglia. METHODS: Here, we used 2-dimensional cultures of human iPSC-derived microglia and transplantation of these cells into the mouse brain to assess the cell-autonomous effects of the A53T mutation on human microglia. RESULTS: We found that A53T-mutant human microglia had an intrinsically increased propensity towards pro-inflammatory activation upon inflammatory stimulus. Additionally, transplanted A53T mutant microglia showed a strong decrease in catalase expression in non-inflammatory conditions, and increased oxidative stress. CONCLUSIONS: Our results indicate that A53T mutant human microglia display cell-autonomous phenotypes that may worsen neuronal damage in early-onset PD.

2.
Biol Psychiatry ; 93(1): 71-81, 2023 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-36372569

RESUMEN

BACKGROUND: Fragile X syndrome (FXS) is characterized by physical abnormalities, anxiety, intellectual disability, hyperactivity, autistic behaviors, and seizures. Abnormal neuronal development in FXS is poorly understood. Data on patients with FXS remain scarce, and FXS animal models have failed to yield successful therapies. In vitro models do not fully recapitulate the morphology and function of human neurons. METHODS: To mimic human neuron development in vivo, we coinjected neural precursor cells derived from FXS patient-derived induced pluripotent stem cells and neural precursor cells derived from corrected isogenic control induced pluripotent stem cells into the brain of neonatal immune-deprived mice. RESULTS: The transplanted cells populated the brain and a proportion differentiated into neurons and glial cells. Immunofluorescence and single and bulk RNA sequencing analyses showed accelerated maturation of FXS neurons after an initial delay. Additionally, we found increased percentages of Arc- and Egr-1-positive FXS neurons and wider dendritic protrusions of mature FXS striatal medium spiny neurons. CONCLUSIONS: This transplantation approach provides new insights into the alterations of neuronal development in FXS by facilitating physiological development of cells in a 3-dimensional context.


Asunto(s)
Síndrome del Cromosoma X Frágil , Células-Madre Neurales , Humanos , Ratones , Animales , Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Células-Madre Neurales/metabolismo , Neuronas/metabolismo , Fenotipo , Encéfalo/metabolismo , Ratones Noqueados
3.
Mol Cell ; 41(1): 93-106, 2011 Jan 07.
Artículo en Inglés | MEDLINE | ID: mdl-21211726

RESUMEN

Protein quality control (PQC) degradation systems protect the cell from the toxic accumulation of misfolded proteins. Because any protein can become misfolded, these systems must be able to distinguish abnormal proteins from normal ones, yet be capable of recognizing the wide variety of distinctly shaped misfolded proteins they are likely to encounter. How individual PQC degradation systems accomplish this remains an open question. Here we show that the yeast nuclear PQC ubiquitin ligase San1 directly recognizes its misfolded substrates via intrinsically disordered N- and C-terminal domains. These disordered domains are punctuated with small segments of order and high sequence conservation that serve as substrate-recognition sites San1 uses to target its different substrates. We propose that these substrate-recognition sites, interspersed among flexible, disordered regions, provide San1 an inherent plasticity which allows it to bind its many, differently shaped misfolded substrates.


Asunto(s)
Pliegue de Proteína , Complejos de Ubiquitina-Proteína Ligasa/fisiología , Secuencia de Aminoácidos , Datos de Secuencia Molecular , Mapeo de Interacción de Proteínas , Estructura Terciaria de Proteína , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/metabolismo , Alineación de Secuencia , Especificidad por Sustrato , Complejos de Ubiquitina-Proteína Ligasa/química , Ubiquitina-Proteína Ligasas/química , Ubiquitina-Proteína Ligasas/metabolismo
4.
PLoS Genet ; 3(12): e219, 2007 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-18069898

RESUMEN

The cyclin-dependent kinase inhibitor p27(KIP1) is a tumor suppressor gene in mice, and loss of p27 protein is a negative prognostic indicator in human cancers. Unlike other tumor suppressors, the p27 gene is rarely mutated in tumors. Therefore misregulation of p27, rather than loss of the gene, is responsible for tumor-associated decreases in p27 protein levels. We performed a functional genomic screen in p27(+/-) mice to identify genes that regulate p27 during lymphomagenesis. This study demonstrated that decreased p27 expression in tumors resulted from altered transcription of the p27 gene, and the retroviral tagging strategy enabled us to pinpoint relevant transcription factors. inhibitor of DNA binding 3 (Id3) was isolated and validated as a transcriptional repressor of p27. We further demonstrated that p27 was a downstream target of Id3 in src-family kinase Lck-driven thymic lymphomagenesis and that p27 was an essential regulator of Lck-dependent thymic maturation during normal T-cell development. Thus, we have identified and characterized transcriptional repression of p27 by Id3 as a new mechanism decreasing p27 protein in tumors.


Asunto(s)
Inhibidor p27 de las Quinasas Dependientes de la Ciclina/genética , Linfoma/genética , Animales , Secuencia de Bases , Diferenciación Celular , Línea Celular Tumoral , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/deficiencia , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Proteínas Inhibidoras de la Diferenciación/genética , Proteínas Inhibidoras de la Diferenciación/metabolismo , Proteína Tirosina Quinasa p56(lck) Específica de Linfocito/genética , Proteína Tirosina Quinasa p56(lck) Específica de Linfocito/metabolismo , Linfoma/metabolismo , Linfoma/virología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Mutantes , Virus de la Leucemia Murina de Moloney/genética , Virus de la Leucemia Murina de Moloney/patogenicidad , Regiones Promotoras Genéticas , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Neoplásico/genética , ARN Neoplásico/metabolismo , ARN Interferente Pequeño/genética , Linfocitos T/citología , Linfocitos T/metabolismo , Transcripción Genética
5.
Proc Natl Acad Sci U S A ; 103(38): 14009-14, 2006 Sep 19.
Artículo en Inglés | MEDLINE | ID: mdl-16966613

RESUMEN

Decreased expression of the CDK inhibitor p27kip1 in human tumors directly correlates with increased resistance to chemotherapies, increased rates of metastasis, and an overall increased rate of patient mortality. It is thought that decreased p27 expression in tumors is caused by increased proteasomal turnover, in particular activation of the pathway governed by the SCFskp2 E3 ubiquitin protein ligase. We have directly tested the importance of the SCFskp-mediated degradation of p27 in tumorigenesis by analyzing the tumor susceptibility of mice that express a form of p27 that cannot be ubiquitinated and degraded by this pathway (p27T187A). In mouse models of both lung and colon cancer down-regulation of p27 promotes tumorigenesis. However, we found that preventing p27 degradation by the SCFskp2 pathway had no impact on tumor incidence or overall survival in either tumor model. Our study unveiled a previously unrecognized role for the control of p27 mRNA abundance in the development of non-small cell lung cancers. In the colon cancer model, the frequency of intestinal adenomas was similarly unaffected by the p27T187A mutation, but, unexpectedly, we found that it inhibited progression of intestinal adenomas to carcinomas. These studies may guide the choice of clinical settings in which pharmacologic inhibitors of the Skp2 pathway might be of therapeutic value.


Asunto(s)
Neoplasias del Colon/metabolismo , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Neoplasias Pulmonares/metabolismo , Inhibidores de Proteínas Quinasas/metabolismo , Proteínas Quinasas Asociadas a Fase-S/metabolismo , Transducción de Señal/fisiología , Animales , Transformación Celular Neoplásica , Neoplasias del Colon/patología , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/genética , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Genes ras , Humanos , Neoplasias Pulmonares/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , ARN Mensajero/metabolismo , Tasa de Supervivencia , Ubiquitina/metabolismo
6.
Curr Biol ; 13(23): 2025-36, 2003 Dec 02.
Artículo en Inglés | MEDLINE | ID: mdl-14653991

RESUMEN

BACKGROUND: Sister chromatid cohesion is needed for proper alignment and segregation of chromosomes during cell division. Chromatids are linked by the multiprotein cohesin complex, which binds to DNA during G(1) and then establishes cohesion during S phase DNA replication. However, many aspects of the mechanisms that establish and maintain cohesion during mitosis remain unclear. RESULTS: We found that mutations in two evolutionarily conserved Drosophila genes, san (separation anxiety) and deco (Drosophila eco1), disrupt centromeric sister chromatid cohesion very early in division. This failure of sister chromatid cohesion does not require separase and is correlated with a failure of the cohesin component Scc1 to accumulate in centromeric regions. It thus appears that these mutations interfere with the establishment of centromeric sister chromatid cohesion. Secondary consequences of these mutations include activation of the spindle checkpoint, causing metaphase delay or arrest. Some cells eventually escape the block but incur many errors in anaphase chromosome segregation. Both san and deco are predicted to encode acetyltransferases, which transfer acetyl groups either to internal lysine residues or to the N terminus of other proteins. The San protein is itself acetylated, and it associates with the Nat1 and Ard1 subunits of the NatA acetyltransferase. CONCLUSIONS: At least two diverse acetyltransferases play vital roles in regulating sister chromatid cohesion during Drosophila mitosis.


Asunto(s)
Acetiltransferasas/genética , Cromátides/metabolismo , Proteínas de Drosophila/genética , Drosophila/fisiología , Mitosis/fisiología , Acetiltransferasas/metabolismo , Animales , Western Blotting , Proteínas de Ciclo Celular/metabolismo , Cromatografía de Afinidad , Proteínas Cromosómicas no Histona , Drosophila/genética , Drosophila/metabolismo , Microscopía Fluorescente , Mutación/fisiología , Proteínas Nucleares , Fosfoproteínas , Proteínas de Saccharomyces cerevisiae , Huso Acromático/fisiología
7.
Development ; 129(20): 4661-75, 2002 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-12361959

RESUMEN

Previous genetic studies indicated intersex (ix) functions only in females and that it acts near the end of the sex determination hierarchy to control somatic sexual differentiation in Drosophila melanogaster. We have cloned ix and characterized its function genetically, molecularly and biochemically. The ix pre-mRNA is not spliced, and ix mRNA is produced in both sexes. The ix gene encodes a 188 amino acid protein, which has a sequence similar to mammalian proteins thought to function as transcriptional activators, and a Caenorhabditis elegans protein that is thought to function as a transcription factor. Bringing together the facts that (1) the ix phenotype is female-specific and (2) functions at the end of the sex determination hierarchy, yet (3) is expressed sex non-specifically and appears likely to encode a transcription factor with no known DNA-binding domain, leads to the inference that ix may require the female-specific protein product of the doublesex (dsx) gene in order to function. Consistent with this inference, we find that for all sexually dimorphic cuticular structures examined, ix and dsx are dependent on each other to promote female differentiation. This dependent relationship also holds for the only known direct target of dsx, the Yolk protein (Yp) genes. Using yeast 2-hybrid assay, immunoprecipitation of recombinant tagged IX and DSX proteins from Drosophila S2 cell extracts, and gel shifts with the tagged IX and DSX(F) proteins, we demonstrate that IX interacts with DSX(F), but not DSX(M). Taken together, the above findings strongly suggest that IX and DSX(F) function in a complex, in which IX acts as a transcriptional co-factor for the DNA-binding DSX(F).


Asunto(s)
Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/crecimiento & desarrollo , Drosophila melanogaster/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Vagina/crecimiento & desarrollo , Empalme Alternativo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , Proteínas de Unión al ADN/genética , Femenino , Regulación del Desarrollo de la Expresión Génica , Masculino , Datos de Secuencia Molecular , Mutación , ARN Mensajero/metabolismo , Diferenciación Sexual/genética , Vitelogeninas
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