Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 21.110
Filtrar
1.
J Med Microbiol ; 70(4)2021 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-33830911

RESUMEN

Introduction. Antipathogenic or antivirulence strategy is to target a virulence pathway that is dispensable for growth, in the hope to mitigate the selection for drug resistance.Hypothesis/Gap Statment. Peroxide stress responses are one of the conserved virulence pathways in bacterial pathogens and thus good targets for antipathogenic strategy.Aim. This study aims to identify a new chemical compound that targets OxyR, the peroxide sensor required for the full virulence of the opportunistic human pathogen, Pseudomonas aeruginosa.Methodology. Computer-based virtual screening under consideration of the 'eNTRy' rules and molecular docking were conducted on the reduced form of the OxyR regulatory domain (RD). Selected hits were validated by their ability to phenocopy the oxyR null mutant and modulate the redox cycle of OxyR.Results. We first isolated three robust chemical hits that inhibit OxyR without affecting prototrophic growth or viability. One (compound 1) of those affected the redox cycle of OxyR in response to H2O2 treatment, in a way to impair its function. Compound 1 displayed selective antibacterial efficacy against P. aeruginosa in Drosophila infection model, without antibacterial activity against Staphylococcus aureus.Conclusion. These results suggest that compound 1 could be an antipathogenic hit inhibiting the P. aeruginosa OxyR. More importantly, our study provides an insight into the computer-based discovery of new-paradigm selective antibacterials to treat Gram-negative bacterial infections presumably with few concerns of drug resistance.


Asunto(s)
Antibacterianos/farmacología , Pseudomonas aeruginosa/efectos de los fármacos , Pseudomonas aeruginosa/patogenicidad , Transactivadores/antagonistas & inhibidores , Animales , Drosophila , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Peróxido de Hidrógeno/farmacología , Simulación del Acoplamiento Molecular , Mutación , Oxidación-Reducción , Estrés Oxidativo/efectos de los fármacos , Infecciones por Pseudomonas/tratamiento farmacológico , Infecciones por Pseudomonas/mortalidad , Pseudomonas aeruginosa/genética , Tasa de Supervivencia , Transactivadores/química , Transactivadores/genética , Transactivadores/metabolismo , Virulencia/efectos de los fármacos , Virulencia/genética
2.
Nat Commun ; 12(1): 2397, 2021 04 23.
Artículo en Inglés | MEDLINE | ID: mdl-33893274

RESUMEN

Gene targeting studies in primary human islets could advance our understanding of mechanisms driving diabetes pathogenesis. Here, we demonstrate successful genome editing in primary human islets using clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 9 (Cas9). CRISPR-based targeting efficiently mutated protein-coding exons, resulting in acute loss of islet ß-cell regulators, like the transcription factor PDX1 and the KATP channel subunit KIR6.2, accompanied by impaired ß-cell regulation and function. CRISPR targeting of non-coding DNA harboring type 2 diabetes (T2D) risk variants revealed changes in ABCC8, SIX2 and SIX3 expression, and impaired ß-cell function, thereby linking regulatory elements in these target genes to T2D genetic susceptibility. Advances here establish a paradigm for genetic studies in human islet cells, and reveal regulatory and genetic mechanisms linking non-coding variants to human diabetes risk.


Asunto(s)
Sistemas CRISPR-Cas , Edición Génica/métodos , Células Secretoras de Insulina/metabolismo , Islotes Pancreáticos/metabolismo , Modelos Genéticos , Secuencia de Bases , Diabetes Mellitus Tipo 2/genética , Regulación de la Expresión Génica , Proteínas de Homeodominio/genética , Humanos , Células Secretoras de Insulina/citología , Islotes Pancreáticos/citología , Canales de Potasio de Rectificación Interna/genética , Transactivadores/genética
3.
Nat Commun ; 12(1): 2340, 2021 04 20.
Artículo en Inglés | MEDLINE | ID: mdl-33879786

RESUMEN

Cancer is characterized by pervasive epigenetic alterations with enhancer dysfunction orchestrating the aberrant cancer transcriptional programs and transcriptional dependencies. Here, we epigenetically characterize human colorectal cancer (CRC) using de novo chromatin state discovery on a library of different patient-derived organoids. By exploring this resource, we unveil a tumor-specific deregulated enhancerome that is cancer cell-intrinsic and independent of interpatient heterogeneity. We show that the transcriptional coactivators YAP/TAZ act as key regulators of the conserved CRC gained enhancers. The same YAP/TAZ-bound enhancers display active chromatin profiles across diverse human tumors, highlighting a pan-cancer epigenetic rewiring which at single-cell level distinguishes malignant from normal cell populations. YAP/TAZ inhibition in established tumor organoids causes extensive cell death unveiling their essential role in tumor maintenance. This work indicates a common layer of YAP/TAZ-fueled enhancer reprogramming that is key for the cancer cell state and can be exploited for the development of improved therapeutic avenues.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Neoplasias Colorrectales/genética , Elementos de Facilitación Genéticos , Epigénesis Genética , Transactivadores/genética , Factores de Transcripción/genética , Regulación Neoplásica de la Expresión Génica , Código de Histonas , Humanos , Modelos Genéticos , Organoides/metabolismo , RNA-Seq , Análisis de la Célula Individual , Células Tumorales Cultivadas
4.
Nat Commun ; 12(1): 1846, 2021 03 23.
Artículo en Inglés | MEDLINE | ID: mdl-33758180

RESUMEN

A wide repertoire of genetic switches has accelerated prokaryotic synthetic biology, while eukaryotic synthetic biology has lagged in the model organism Saccharomyces cerevisiae. Eukaryotic genetic switches are larger and more complex than prokaryotic ones, complicating the rational design and evolution of them. Here, we present a robust workflow for the creation and evolution of yeast genetic switches. The selector system was designed so that both ON- and OFF-state selection of genetic switches is completed solely by liquid handling, and it enabled parallel screen/selection of different motifs with different selection conditions. Because selection threshold of both ON- and OFF-state selection can be flexibly tuned, the desired selection conditions can be rapidly pinned down for individual directed evolution experiments without a prior knowledge either on the library population. The system's utility was demonstrated using 20 independent directed evolution experiments, yielding genetic switches with elevated inducer sensitivities, inverted switching behaviours, sensory functions, and improved signal-to-noise ratio (>100-fold induction). The resulting yeast genetic switches were readily integrated, in a plug-and-play manner, into an AND-gated carotenoid biosynthesis pathway.


Asunto(s)
Evolución Molecular Dirigida/métodos , Genes de Cambio , Ingeniería Genética/métodos , Técnicas Genéticas , Saccharomyces cerevisiae/genética , Biología Sintética/métodos , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Basidiomycota/genética , Basidiomycota/metabolismo , Citometría de Flujo , Biblioteca de Genes , Genes Reporteros , Floroglucinol/análogos & derivados , Floroglucinol/farmacología , Regiones Promotoras Genéticas , Proteínas Represoras/química , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Relación Señal-Ruido , Tetraciclina/farmacología , Transactivadores/química , Transactivadores/genética , Transactivadores/metabolismo , beta Caroteno/biosíntesis , beta Caroteno/genética , beta Caroteno/metabolismo
5.
Int J Mol Sci ; 22(4)2021 Feb 13.
Artículo en Inglés | MEDLINE | ID: mdl-33668437

RESUMEN

Developmental growth and patterning are regulated by an interconnected signalling network of several pathways. In Drosophila, the Warts (Wts) kinase, a component of the Hippo signalling pathway, plays an essential role in regulating transcription and growth by phosphorylating its substrate Yorkie (Yki). The phosphorylation of Yki critically influences its localisation and activity as a transcriptional coactivator. In this study, we identified the homeodomain-interacting protein kinase (Hipk) as another kinase that phosphorylates Yki and mapped several sites of Yki phosphorylated by Hipk, using in vitro analysis: Ser168, Ser169/Ser172 and Ser255. These sites might provide auxiliary input for Yki regulation in vivo, as transgenic flies with mutations in these show prominent phenotypes; Hipk, therefore, represents an additional upstream regulator of Yki that works in concert with Wts.


Asunto(s)
Proteínas de Drosophila/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal , Transactivadores/metabolismo , Animales , Proteínas de Drosophila/genética , Drosophila melanogaster , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Nucleares/genética , Fosforilación/genética , Proteínas Quinasas/genética , Proteínas Serina-Treonina Quinasas/genética , Transactivadores/genética
6.
Molecules ; 26(5)2021 Feb 26.
Artículo en Inglés | MEDLINE | ID: mdl-33652602

RESUMEN

Hepatitis B virus (HBV) is a circular, and partially double-stranded DNA virus. Upon infection, the viral genome is translocated into the cell nucleus, generating the covalently closed circular DNA (cccDNA) intermediate, and forming a mini chromosome. HBV HBx is a small protein displaying multiple roles in HBV-infected cells, and in different subcellular locations. In the nucleus, the HBx protein is required to initiate and maintain viral transcription from the viral mini chromosome. In contrast, HBx also functions in the cytoplasm, where it is able to alter multiple cellular functions such as mitochondria metabolism, apoptosis and signal transduction pathways. It has been reported that in cultured cells, at low expression levels, the HBx protein is localized in the nucleus, whereas at high expression levels, it accumulates in the cytoplasm. This dynamic subcellular distribution of HBx might be essential to exert its multiple roles during viral infection. However, the mechanism that regulates different subcellular localizations of the HBx protein is unknown. We have previously taken a bioinformatics approach to investigate whether HBx might be regulated via post-translational modification, and we have proposed that the multiple nucleocytoplasmic functions of HBx might be regulated by an evolutionarily conserved mechanism via phosphorylation. In the current study, phylogenetically conserved amino acids of HBx with a high potential of phosphorylation were targeted for site-directed mutagenesis. Two conserved serine (Ser25 and Ser41), and one conserved threonine (Thr81) amino acids were replaced by either alanine or aspartic acid residues to simulate an unphosphorylated or phosphorylated state, respectively. Human hepatoma cells were transfected with increasing amounts of the HBx DNA constructs, and the cells were analyzed by fluorescence microscopy. Together, our results show that the nucleocytoplasmic distribution of the HBx protein could be regulated by phosphorylation since some of the modified proteins were mainly confined to distinct subcellular compartments. Remarkably, both HBx Ser41A, and HBx Thr81D proteins were predominantly localized within the nuclear compartment throughout the different expression levels of HBx mutants.


Asunto(s)
Carcinoma Hepatocelular/genética , Hepatitis B/genética , Neoplasias Hepáticas/genética , Transactivadores/genética , Proteínas Reguladoras y Accesorias Virales/genética , Secuencia de Aminoácidos/genética , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/virología , Secuencia Conservada/genética , Regulación Viral de la Expresión Génica/genética , Genoma Viral/genética , Células Hep G2 , Hepatitis B/patología , Hepatitis B/virología , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/patogenicidad , Humanos , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/virología , Fosforilación/genética , Filogenia
7.
Mol Med Rep ; 23(6)2021 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-33786620

RESUMEN

Chromosome 14 open reading frame 166 (C14orf166) encodes a 28­kDa nuclear and cytoplasmic protein that is involved in viral infection, RNA metabolism, and centrosome structure. It binds to the polymerase acidic protein subunit of the influenza A virus, which is associated with several transcription factors, RNA polymerase II, to activate transcription initiation and promote virus infection. It also interacts with a mature hepatitis C virus core protein to regulate the infection process. In physiological conditions, C14orf166 associates with the proteins, DDX1, HSPC117 and FAM98B, and regulates RNA metabolism and fate. In addition, C14orf166 is overexpressed in a variety of cancer types. Upregulation of C14orf166 may contribute toward cancer malignancy through its impact on glycogen synthase kinase 3ß­mediated signaling, the downregulation of retinoblastoma protein, or the upregulation of IL­6. Therefore, C14orf166 could be used as a biomarker for the diagnosis and prognosis of various cancer types. This review summarized the existent literature about C14orf166, focusing on its functions in physiological and pathological situations.


Asunto(s)
Neoplasias/genética , ARN/metabolismo , Transactivadores/genética , Virosis/genética , Animales , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Humanos , Neoplasias/metabolismo , ARN/genética , Transactivadores/metabolismo , Virosis/metabolismo
8.
Science ; 371(6536): 1350-1355, 2021 03 26.
Artículo en Inglés | MEDLINE | ID: mdl-33632892

RESUMEN

Mitogens trigger cell division in animals. In plants, cytokinins, a group of phytohormones derived from adenine, stimulate cell proliferation. Cytokinin signaling is initiated by membrane-associated histidine kinase receptors and transduced through a phosphorelay system. We show that in the Arabidopsis shoot apical meristem (SAM), cytokinin regulates cell division by promoting nuclear shuttling of Myb-domain protein 3R4 (MYB3R4), a transcription factor that activates mitotic gene expression. Newly synthesized MYB3R4 protein resides predominantly in the cytoplasm. At the G2-to-M transition, rapid nuclear accumulation of MYB3R4-consistent with an associated transient peak in cytokinin concentration-feeds a positive feedback loop involving importins and initiates a transcriptional cascade that drives mitosis and cytokinesis. An engineered nuclear-restricted MYB3R4 mimics the cytokinin effects of enhanced cell proliferation and meristem growth.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/citología , Arabidopsis/metabolismo , División Celular , Citocininas/metabolismo , Transactivadores/metabolismo , Transporte Activo de Núcleo Celular , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Puntos de Control del Ciclo Celular , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Regulación de la Expresión Génica de las Plantas , Carioferinas/metabolismo , Meristema/metabolismo , Mitosis/genética , Reguladores del Crecimiento de las Plantas/metabolismo , Transducción de Señal , Transactivadores/genética
9.
Nucleic Acids Res ; 49(5): 2569-2582, 2021 03 18.
Artículo en Inglés | MEDLINE | ID: mdl-33621320

RESUMEN

During oogenesis, oocytes gain competence and subsequently undergo meiotic maturation and prepare for embryonic development; trimethylated histone H3 on lysine-4 (H3K4me3) mediates a wide range of nuclear events during these processes. Oocyte-specific knockout of CxxC-finger protein 1 (CXXC1, also known as CFP1) impairs H3K4me3 accumulation and causes changes in chromatin configurations. This study investigated the changes in genomic H3K4me3 landscapes in oocytes with Cxxc1 knockout and the effects on other epigenetic factors such as the DNA methylation, H3K27me3, H2AK119ub1 and H3K36me3. H3K4me3 is overall decreased after knocking out Cxxc1, including both the promoter region and the gene body. CXXC1 and MLL2, which is another histone H3 methyltransferase, have nonoverlapping roles in mediating H3K4 trimethylation during oogenesis. Cxxc1 deletion caused a decrease in DNA methylation levels and affected H3K27me3 and H2AK119ub1 distributions, particularly at regions with high DNA methylation levels. The changes in epigenetic networks implicated by Cxxc1 deletion were correlated with the transcriptional changes in genes in the corresponding genomic regions. This study elucidates the epigenetic changes underlying the phenotypes and molecular defects in oocytes with deleted Cxxc1 and highlights the role of CXXC1 in orchestrating multiple factors that are involved in establishing the appropriate epigenetic states of maternal genome.


Asunto(s)
Epigénesis Genética , Oocitos/metabolismo , Transactivadores/fisiología , Animales , Células Cultivadas , Metilación de ADN , Femenino , Eliminación de Gen , Genoma , Código de Histonas , Histonas/metabolismo , Ratones , Ratones Endogámicos C57BL , Regiones Promotoras Genéticas , Transactivadores/genética , Transcripción Genética
10.
Int J Mol Sci ; 22(3)2021 Jan 22.
Artículo en Inglés | MEDLINE | ID: mdl-33499042

RESUMEN

In this review, we discuss the major histocompatibility complex (MHC) class II transactivator (CIITA), which is the master regulator of MHC class II gene expression. CIITA is the founding member of the mammalian nucleotide-binding and leucine-rich-repeat (NLR) protein family but stood apart for a long time as the only transcriptional regulator. More recently, it was found that its closest homolog, NLRC5 (NLR protein caspase activation and recruitment domain (CARD)-containing 5), is a regulator of MHC-I gene expression. Both act as non-DNA-binding activators through multiple protein-protein interactions with an MHC enhanceosome complex that binds cooperatively to a highly conserved combinatorial cis-acting module. Thus, the regulation of MHC-II expression is regulated largely through the differential expression of CIITA. In addition to the well-defined role of CIITA in MHC-II GENE regulation, we will discuss several other aspects of CIITA functions, such as its role in cancer, its role as a viral restriction element contributing to intrinsic immunity, and lastly, its very recently discovered role as an inhibitor of Ebola and SARS-Cov-2 virus replication. We will briefly touch upon the recently discovered role of NLRP3 as a transcriptional regulator, which suggests that transcriptional regulation is, after all, not such an unusual feature for NLR proteins.


Asunto(s)
Genes MHC Clase II , Proteínas NLR/metabolismo , Proteínas Nucleares/metabolismo , Transactivadores/metabolismo , Animales , /metabolismo , Ebolavirus/fisiología , Regulación de la Expresión Génica , Fiebre Hemorrágica Ebola/genética , Fiebre Hemorrágica Ebola/metabolismo , Humanos , Proteínas NLR/genética , Neoplasias/genética , Neoplasias/metabolismo , Proteínas Nucleares/genética , Mapas de Interacción de Proteínas , Transactivadores/genética , Replicación Viral
11.
Nat Commun ; 12(1): 383, 2021 01 15.
Artículo en Inglés | MEDLINE | ID: mdl-33452256

RESUMEN

The transcription factor p63 mediates distinct cellular responses, primarily regulating epithelial and oocyte biology. In addition to the two amino terminal isoforms, TAp63 and ΔNp63, the 3'-end of p63 mRNA undergoes tissue-specific alternative splicing that leads to several isoforms, including p63α, p63ß and p63γ. To investigate in vivo how the different isoforms fulfil distinct functions at the cellular and developmental levels, we developed a mouse model replacing the p63α with p63ß by deletion of exon 13 in the Trp63 gene. Here, we report that whereas in two organs physiologically expressing p63α, such as thymus and skin, no abnormalities are detected, total infertility is evident in heterozygous female mice. A sharp reduction in the number of primary oocytes during the first week after birth occurs as a consequence of the enhanced expression of the pro-apoptotic transcriptional targets Puma and Noxa by the tetrameric, constitutively active, TAp63ß isoform. Hence, these mice show a condition of ovary dysfunction, resembling human primary ovary insufficiency. Our results show that the p63 C-terminus is essential in TAp63α-expressing primary oocytes to control cell death in vivo, expanding the current understanding of human primary ovarian insufficiency.


Asunto(s)
Infertilidad Femenina/genética , Oocitos/patología , Insuficiencia Ovárica Primaria/genética , Transactivadores/genética , Empalme Alternativo/genética , Animales , Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/genética , Células Cultivadas , Modelos Animales de Enfermedad , Exones/genética , Femenino , Heterocigoto , Humanos , Infertilidad Femenina/patología , Masculino , Ratones , Mutación , Cultivo Primario de Células , Insuficiencia Ovárica Primaria/complicaciones , Insuficiencia Ovárica Primaria/patología , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/genética , Transactivadores/metabolismo , Activación Transcripcional , Proteínas Supresoras de Tumor/genética
12.
Nat Commun ; 12(1): 410, 2021 01 18.
Artículo en Inglés | MEDLINE | ID: mdl-33462227

RESUMEN

Active DNA demethylation is required for sexual reproduction in plants but the molecular determinants underlying this epigenetic control are not known. Here, we show in Arabidopsis thaliana that the DNA glycosylases DEMETER (DME) and REPRESSOR OF SILENCING 1 (ROS1) act semi-redundantly in the vegetative cell of pollen to demethylate DNA and ensure proper pollen tube progression. Moreover, we identify six pollen-specific genes with increased DNA methylation as well as reduced expression in dme and dme;ros1. We further show that for four of these genes, reinstalling their expression individually in mutant pollen is sufficient to improve male fertility. Our findings demonstrate an essential role of active DNA demethylation in regulating genes involved in pollen function.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiología , Desmetilación del ADN , Regulación de la Expresión Génica de las Plantas , N-Glicosil Hidrolasas/metabolismo , Proteínas Nucleares/metabolismo , Transactivadores/metabolismo , Proteínas de Arabidopsis/genética , Fertilidad/genética , Regulación del Desarrollo de la Expresión Génica , Mutación , N-Glicosil Hidrolasas/genética , Proteínas Nucleares/genética , Plantas Modificadas Genéticamente , Tubo Polínico/crecimiento & desarrollo , Transactivadores/genética
13.
Nucleic Acids Res ; 49(2): 832-846, 2021 01 25.
Artículo en Inglés | MEDLINE | ID: mdl-33406256

RESUMEN

The Salmonella genomic island 1 (SGI1) and its variants are mobilized by IncA and IncC conjugative plasmids. SGI1-family elements and their helper plasmids are effective transporters of multidrug resistance determinants. SGI1 exploits the transfer apparatus of the helper plasmid and hijacks its activator complex, AcaCD, to trigger the expression of several SGI1 genes. In this way, SGI1 times its excision from the chromosome to the helper entry and expresses mating pore components that enhance SGI1 transfer. The SGI1-encoded T4SS components and the FlhDC-family activator proved to be interchangeable with their IncC-encoded homologs, indicating multiple interactions between SGI1 and its helpers. As a new aspect of this crosstalk, we report here the helper-induced replication of SGI1, which requires both activators, AcaCD and FlhDCSGI1, and significantly increases the stability of SGI1 when coexists with the helper plasmid. We have identified the oriVSGI1 and shown that S004-repA operon encodes for a translationally coupled leader protein and an IncN2/N3-related RepA that are expressed under the control of the AcaCD-responsive promoter PS004. This replicon transiently maintains SGI1 as a 4-8-copy plasmid, not only stabilizing the island but also contributing to the fast displacement of the helper plasmid.


Asunto(s)
Proteínas Bacterianas/genética , Cromosomas Bacterianos/genética , Conjugación Genética/genética , Farmacorresistencia Bacteriana Múltiple/genética , Secuencias Repetitivas Esparcidas/genética , Salmonella typhimurium/genética , Proteínas Bacterianas/metabolismo , ADN Helicasas/genética , ADN Helicasas/metabolismo , Replicación del ADN , Dosificación de Gen , Regulación Bacteriana de la Expresión Génica/genética , Genes Reporteros , Integrasas/metabolismo , Operón/genética , Filogenia , Plásmidos/genética , Regiones Promotoras Genéticas/genética , Biosíntesis de Proteínas , Recombinasas/metabolismo , Replicón/genética , Alineación de Secuencia , Transactivadores/genética , Transactivadores/metabolismo
14.
Nat Genet ; 53(2): 166-173, 2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33462483

RESUMEN

Despite the strong genetic basis of psychiatric disorders, the underlying molecular mechanisms are largely unmapped. RNA-binding proteins (RBPs) are responsible for most post-transcriptional regulation, from splicing to translation to localization. RBPs thus act as key gatekeepers of cellular homeostasis, especially in the brain. However, quantifying the pathogenic contribution of noncoding variants impacting RBP target sites is challenging. Here, we leverage a deep learning approach that can accurately predict the RBP target site dysregulation effects of mutations and discover that RBP dysregulation is a principal contributor to psychiatric disorder risk. RBP dysregulation explains a substantial amount of heritability not captured by large-scale molecular quantitative trait loci studies and has a stronger impact than common coding region variants. We share the genome-wide profiles of RBP dysregulation, which we use to identify DDHD2 as a candidate schizophrenia risk gene. This resource provides a new analytical framework to connect the full range of RNA regulation to complex disease.


Asunto(s)
Trastornos Mentales/genética , Fosfolipasas/genética , Proteínas de Unión al ARN/genética , Regiones no Traducidas 3' , Aprendizaje Profundo , Regulación de la Expresión Génica , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo , Humanos , Mutación , Proteínas del Factor Nuclear 90/genética , Factores de Elongación de Péptidos/genética , Polimorfismo de Nucleótido Simple , Sitios de Carácter Cuantitativo , ARN Helicasas/genética , Procesamiento Postranscripcional del ARN , Ribonucleoproteína Nuclear Pequeña U5/genética , Esquizofrenia/genética , Transactivadores/genética
15.
Mol Cell ; 81(3): 571-583.e6, 2021 02 04.
Artículo en Inglés | MEDLINE | ID: mdl-33412111

RESUMEN

The arms race between bacteria and phages has led to the evolution of diverse anti-phage defenses, several of which are controlled by quorum-sensing pathways. In this work, we characterize a quorum-sensing anti-activator protein, Aqs1, found in Pseudomonas phage DMS3. We show that Aqs1 inhibits LasR, the master regulator of quorum sensing, and present the crystal structure of the Aqs1-LasR complex. The 69-residue Aqs1 protein also inhibits PilB, the type IV pilus assembly ATPase protein, which blocks superinfection by phages that require the pilus for infection. This study highlights the remarkable ability of small phage proteins to bind multiple host proteins and disrupt key biological pathways. As quorum sensing influences various anti-phage defenses, Aqs1 provides a mechanism by which infecting phages might simultaneously dampen multiple defenses. Because quorum-sensing systems are broadly distributed across bacteria, this mechanism of phage counter-defense may play an important role in phage-host evolutionary dynamics.


Asunto(s)
Proteínas Bacterianas/metabolismo , Bacteriófagos/metabolismo , Pseudomonas aeruginosa/metabolismo , Percepción de Quorum , Transactivadores/metabolismo , Proteínas Virales/metabolismo , Proteínas Bacterianas/genética , Bacteriófagos/genética , Bacteriófagos/patogenicidad , Fimbrias Bacterianas/metabolismo , Interacciones Huésped-Patógeno , Oxidorreductasas/genética , Oxidorreductasas/metabolismo , Unión Proteica , Dominios y Motivos de Interacción de Proteínas , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/crecimiento & desarrollo , Piocianina/metabolismo , Transactivadores/genética , Proteínas Virales/genética
16.
Am J Pathol ; 191(4): 618-630, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33485866

RESUMEN

CD24 is overexpressed in many human cancers and is a driver of tumor progression. Herein, molecular mechanisms leading to up-regulation of CD24 in prostate cancer were studied. DNA methylation of the CD24 gene promoter at four loci using quantitative methylation-specific PCR was evaluated. Expression of CD24 in tumor tissues was studied by immunohistochemistry. To corroborate the results in vitro, ERG-inducible LNCaP TMPRSS2:ERG (T2E) cells and luciferase promoter assays were used. DNA methylation of the CD24 promoter was significantly higher in tumors than in benign tissue and was associated with biochemical recurrence-free survival, tumor grade, and stage. CD24 mRNA and protein expression were significantly higher in T2E-positive, ERG-overexpressing, and/or PTEN-deficient cases. Higher levels of CD24 protein expression conferred shorter biochemical recurrence-free survival, and these observations were confirmed using The Cancer Genome Atlas prostate adenocarcinoma data. In silico analysis of the CD24 promoter revealed an ERG binding site in between the DNA methylation sites. ERG overexpression led to a strong induction of CD24 mRNA and protein expression. Luciferase promoter assays using the wild-type and mutated ERG binding site within the CD24 promoter showed ERG-dependent activation. Collectively, our results suggest that promoter DNA methylation of the CD24 gene and T2E fusion status are factors involved in the up-regulation of CD24 in patients with prostate cancer.


Asunto(s)
Antígeno CD24/metabolismo , ADN/metabolismo , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Regulador Transcripcional ERG/metabolismo , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/genética , Línea Celular Tumoral , Metilación de ADN/fisiología , Humanos , Masculino , Persona de Mediana Edad , Proteínas de Fusión Oncogénica/genética , Proteínas de Fusión Oncogénica/metabolismo , Transactivadores/genética , Regulador Transcripcional ERG/genética
17.
Dev Cell ; 56(1): 81-94.e10, 2021 01 11.
Artículo en Inglés | MEDLINE | ID: mdl-33400912

RESUMEN

Homeostasis of intestinal epithelia is maintained by coordination of the proper rate of regeneration by stem cell division with the rate of cell loss. Regeneration of host epithelia is normally quiescent upon colonization of commensal bacteria; however, the epithelia often develop dysplasia in a context-dependent manner, the cause and underlying mechanism of which remain unclear. Here, we show that in Drosophila intestine, autophagy lowers the sensitivity of differentiated enterocytes to reactive oxygen species (ROS) that are produced in response to commensal bacteria. We find that autophagy deficiency provokes ROS-dependent excessive regeneration and subsequent epithelial dysplasia and barrier dysfunction. Mechanistically, autophagic substrate Ref(2)P/p62, which co-localizes and physically interacts with Dachs, a Hippo signaling regulator, accumulates upon autophagy deficiency and thus inactivates Hippo signaling, resulting in stem cell over-proliferation non-cell autonomously. Our findings uncover a mechanism whereby suppression of undesirable regeneration by autophagy maintains long-term homeostasis of intestinal epithelia.


Asunto(s)
Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila/metabolismo , Mucosa Intestinal/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Células Madre/metabolismo , Transactivadores/metabolismo , Envejecimiento/metabolismo , Animales , Animales Modificados Genéticamente , Autofagia/efectos de los fármacos , Autofagia/genética , Proliferación Celular/genética , Proteínas de Unión al ADN/genética , Sulfato de Dextran/toxicidad , Drosophila/genética , Drosophila/inmunología , Drosophila/fisiología , Proteínas de Drosophila/genética , Enterocitos/metabolismo , Técnicas de Silenciamiento del Gen , Estudio de Asociación del Genoma Completo , Homeostasis , Inmunohistoquímica , Proteínas Inhibidoras de la Apoptosis/genética , Proteínas Inhibidoras de la Apoptosis/metabolismo , Mucosa Intestinal/citología , Mucosa Intestinal/efectos de los fármacos , Intestinos/citología , Intestinos/efectos de los fármacos , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Musculares , Miosinas/genética , Miosinas/metabolismo , Proteínas Nucleares/genética , Proteínas Serina-Treonina Quinasas/genética , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Transactivadores/genética , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo
18.
Science ; 371(6524): 52-57, 2021 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-33384370

RESUMEN

Neuroendocrine (NE) cells are epithelial cells that possess many of the characteristics of neurons, including the presence of secretory vesicles and the ability to sense environmental stimuli. The normal physiologic functions of solitary airway NE cells remain a mystery. We show that mouse and human airway basal stem cells sense hypoxia. Hypoxia triggers the direct differentiation of these stem cells into solitary NE cells. Ablation of these solitary NE cells during hypoxia results in increased epithelial injury, whereas the administration of the NE cell peptide CGRP rescues this excess damage. Thus, we identify stem cells that directly sense hypoxia and respond by differentiating into solitary NE cells that secrete a protective peptide that mitigates hypoxic injury.


Asunto(s)
Diferenciación Celular , Hipoxia/patología , Células Neuroendocrinas/fisiología , Oxígeno/fisiología , Células Madre/fisiología , Tráquea/citología , Anaerobiosis , Animales , Péptido Relacionado con Gen de Calcitonina/metabolismo , Péptido Relacionado con Gen de Calcitonina/farmacología , Proteína Similar al Receptor de Calcitonina/metabolismo , Recuento de Células , Eliminación de Gen , Humanos , Hipoxia/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Ratones , Ratones Mutantes , Células Neuroendocrinas/citología , Prolil Hidroxilasas/metabolismo , Células Madre/citología , Células Madre/efectos de los fármacos , Transactivadores/genética
19.
Appl Environ Microbiol ; 87(6)2021 02 26.
Artículo en Inglés | MEDLINE | ID: mdl-33397700

RESUMEN

Vibrio species, including the squid symbiont Vibrio fischeri, become competent to take up DNA under specific conditions. For example, V. fischeri becomes competent when grown in the presence of chitin oligosaccharides or upon overproduction of the competence regulatory factor TfoX. While little is known about the regulatory pathway(s) that controls V. fischeri competence, this microbe encodes homologs of factors that control competence in the well-studied V. cholerae To further develop V. fischeri as a genetically tractable organism, we evaluated the roles of some of these competence homologs. Using TfoX-overproducing cells, we found that competence depends upon LitR, the homolog of V. cholerae master quorum-sensing and competence regulator HapR, and upon homologs of putative pilus genes that in V. cholerae facilitate DNA uptake. Disruption of genes for negative regulators upstream of LitR, namely, the LuxO protein and the small RNA (sRNA) Qrr1, resulted in increased transformation frequencies. Unlike LitR-controlled light production, however, competence did not vary with cell density under tfoX overexpression conditions. Analogous to the case with V. cholerae, the requirement for LitR could be suppressed by loss of the Dns nuclease. We also found a role for the putative competence regulator CytR. Finally, we determined that transformation frequencies varied depending on the TfoX-encoding plasmid, and we developed a new dual tfoX and litR overexpression construct that substantially increased the transformation frequency of a less genetically tractable strain. By advancing the ease of genetic manipulation of V. fischeri, these findings will facilitate the rapid discovery of genes involved in physiologically relevant processes, such as biofilm formation and host colonization.IMPORTANCE The ability of bacteria to take up DNA (competence) and incorporate foreign DNA into their genomes (transformation) permits them to rapidly evolve and gain new traits and/or acquire antibiotic resistances. It also facilitates laboratory-based investigations into mechanisms of specific phenotypes, such as those involved in host colonization. Vibrio fischeri has long been a model for symbiotic bacterium-host interactions as well as for other aspects of its physiology, such as bioluminescence and biofilm formation. Competence of V. fischeri can be readily induced upon overexpression of the competence factor TfoX. Relatively little is known about the V. fischeri competence pathway, although homologs of factors known to be important in V. cholerae competence exist. By probing the importance of putative competence factors that control transformation of V. fischeri, this work deepens our understanding of the competence process and advances our ability to genetically manipulate this important model organism.


Asunto(s)
Aliivibrio fischeri/genética , Aliivibrio fischeri/metabolismo , Proteínas Bacterianas/genética , ADN/metabolismo , Transactivadores/genética , Transformación Bacteriana
20.
Cell Mol Life Sci ; 78(7): 3525-3542, 2021 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-33469705

RESUMEN

Metastasis Associated in Colon Cancer 1 (MACC1) is a novel prognostic, predictive and causal biomarker for tumor progression and metastasis in many cancer types, including colorectal cancer. Besides its clinical value, little is known about its molecular function. Its similarity to SH3BP4, involved in regulating uptake and recycling of transmembrane receptors, suggests a role of MACC1 in endocytosis. By exploring the MACC1 interactome, we identified the clathrin-mediated endocytosis (CME)-associated proteins CLTC, DNM2 and AP-2 as MACC1 binding partners. We unveiled a MACC1-dependent routing of internalized transferrin receptor towards recycling. Elevated MACC1 expression caused also the activation and internalization of EGFR, a higher rate of receptor recycling, as well as earlier and stronger receptor activation and downstream signaling. These effects are limited by deletion of CME-related protein interaction sites in MACC1. Thus, MACC1 regulates CME and receptor recycling, causing increased growth factor-mediated downstream signaling and cell proliferation. This novel mechanism unveils potential therapeutic intervention points restricting MACC1-driven metastasis.


Asunto(s)
Clatrina/metabolismo , Neoplasias Colorrectales/patología , Endocitosis , Regulación Neoplásica de la Expresión Génica , Receptores de Transferrina/metabolismo , Transactivadores/metabolismo , Animales , Apoptosis , Biomarcadores de Tumor/metabolismo , Proliferación Celular , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Receptores ErbB/genética , Receptores ErbB/metabolismo , Humanos , Ratones , Proteoma/análisis , Proteoma/metabolismo , Receptores de Transferrina/genética , Transactivadores/genética , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA
...