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1.
Artigo em Russo | MEDLINE | ID: mdl-33081454

RESUMO

OBJECTIVE: To evaluate the frequency of C9orf72-associated frontotemporal dementia (FTD) in the Russian population and to study clinical features of GGGGCC-repeat expansion carriers. MATERIAL AND METHODS: Twenty-eight patients with FTD are included in the study: 15 with a behavioral variant of FTD (bvFTD) and 13 with a agrammatic/non-fluent variant of primary progressive aphasia (avPPA). The mean age was 62 years (34-80), the mean disease duration was 4 years (1-10). The positive family history was noted in 46% of cases. DNA diagnosis was performed using repeat-primed polymerase chain reaction. RESULTS: The frequency of the C9orf72 repeat expansion in patients with FTD was 14%, in patients with bvFTD 20%, in patients with avPPA 8%. The mean age of disease onset in the expansion carriers was 63 (55-75) years. The frequency of the C9orf72 repeats expansion in familial FTD cases was 31%, in sporadic cases 7%. bvFTD with parkinsonian syndrome was noted in two out of four cases, bvFTD with amyotrophic lateral sclerosis (ALS) was shown in one case, avPPA with ALS was shown in one case. One female patient with bvFTD with parkinsonian syndrome presented with cognitive fluctuations that required a differential diagnosis with Lewy body disease. CONCLUSION: This is the first study of the genetic structure of FTD in the Russian population. The prevalence and clinical characteristics of C9orf72-associated FTD were defined, in particular, the spectrum of motor symptoms was shown along with behavioral and aphasic disturbances. DNA diagnosis plays an important role in confirming the diagnosis and selection of patients for potential disease-modifying treatment.


Assuntos
Esclerose Amiotrófica Lateral , Demência Frontotemporal , Adulto , Idoso , Idoso de 80 Anos ou mais , Esclerose Amiotrófica Lateral/diagnóstico , Esclerose Amiotrófica Lateral/epidemiologia , Esclerose Amiotrófica Lateral/genética , Proteína C9orf72/genética , Feminino , Demência Frontotemporal/diagnóstico , Demência Frontotemporal/epidemiologia , Demência Frontotemporal/genética , Humanos , Pessoa de Meia-Idade , Proteínas/genética , Federação Russa/epidemiologia
2.
Nat Commun ; 11(1): 4708, 2020 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-32948758

RESUMO

While the field of microbiology has adapted to the study of complex microbiomes via modern meta-omics techniques, we have not updated our basic knowledge regarding the quantitative levels of DNA, RNA and protein molecules within a microbial cell, which ultimately control cellular function. Here we report the temporal measurements of absolute RNA and protein levels per gene within a mixed bacterial-archaeal consortium. Our analysis of this data reveals an absolute protein-to-RNA ratio of 102-104 for bacterial populations and 103-105 for an archaeon, which is more comparable to Eukaryotic representatives' humans and yeast. Furthermore, we use the linearity between the metaproteome and metatranscriptome over time to identify core functional guilds, hence using a fundamental biological feature (i.e., RNA/protein levels) to highlight phenotypical complementarity. Our findings show that upgrading multi-omic toolkits with traditional absolute measurements unlocks the scaling of core biological questions to dynamic and complex microbiomes, creating a deeper insight into inter-organismal relationships that drive the greater community function.


Assuntos
Microbiota/genética , Microbiota/fisiologia , Proteínas/genética , Proteínas/metabolismo , RNA/genética , RNA/metabolismo , Archaea/genética , Archaea/metabolismo , Bactérias/genética , Bactérias/metabolismo , DNA , Perfilação da Expressão Gênica , Genoma Microbiano , Humanos , Metabolômica , Fenótipo , Proteoma , Proteômica , Transcriptoma , Leveduras
3.
Anticancer Res ; 40(10): 5601-5609, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32988884

RESUMO

BACKGROUND/AIM: Since pathways involving LRRC17 are related to the survival of patients with various cancers, we analyzed LRRC17 as a prognostic gene in serous ovarian cancer. MATERIALS AND METHODS: Data were collected from Gene Expression Omnibus (GSE9891, GSE13876, and GSE26712) and The Cancer Genome Atlas (TCGA). We performed survival analyses using C statistics, area under the curve, survival plot with optimal cutoff level, and cox proportional regression. Zebrafish embryos were used as an in vivo model. RESULTS: The prognosis of patients with high LRRC17 expression was poorer than that of patients with low LRRC17 expression. Multivariate regression analysis showed that LRRC17 expression, age, and stage were independently related with survival. Knockdown of lrrc17 reduced survival rate and delayed development in zebrafish embryos. We also found that lrrc17 is important for cell viability by protecting from p53-dependent apoptosis. CONCLUSION: LRRC17 could be a prognostic gene in ovarian cancer as it regulates cancer cell viability through the p53 pathway.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias Ovarianas/genética , Proteínas/genética , Proteína Supressora de Tumor p53/genética , Idoso , Apoptose/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Técnicas de Silenciamento de Genes , Humanos , Pessoa de Meia-Idade , Neoplasias Ovarianas/patologia , Prognóstico
4.
Nat Commun ; 11(1): 4459, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32900997

RESUMO

The origins of multicellular physiology are tied to evolution of gene expression. Genes can shift expression as organisms evolve, but how ancestral expression influences altered descendant expression is not well understood. To examine this, we amalgamate 1,903 RNA-seq datasets from 182 research projects, including 6 organs in 21 vertebrate species. Quality control eliminates project-specific biases, and expression shifts are reconstructed using gene-family-wise phylogenetic Ornstein-Uhlenbeck models. Expression shifts following gene duplication result in more drastic changes in expression properties than shifts without gene duplication. The expression properties are tightly coupled with protein evolutionary rate, depending on whether and how gene duplication occurred. Fluxes in expression patterns among organs are nonrandom, forming modular connections that are reshaped by gene duplication. Thus, if expression shifts, ancestral expression in some organs induces a strong propensity for expression in particular organs in descendants. Regardless of whether the shifts are adaptive or not, this supports a major role for what might be termed preadaptive pathways of gene expression evolution.


Assuntos
Evolução Molecular , Transcriptoma , Animais , Bases de Dados de Ácidos Nucleicos , Feminino , Duplicação Gênica , Humanos , Masculino , Modelos Genéticos , Família Multigênica , Especificidade de Órgãos , Filogenia , Proteínas/genética , RNA-Seq , Especificidade da Espécie , Vertebrados/classificação , Vertebrados/genética
5.
Nat Commun ; 11(1): 4641, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32934213

RESUMO

Despite recent advances in circuit engineering, the design of genetic networks in mammalian cells is still painstakingly slow and fraught with inexplicable failures. Here, we demonstrate that transiently expressed genes in mammalian cells compete for limited transcriptional and translational resources. This competition results in the coupling of otherwise independent exogenous and endogenous genes, creating a divergence between intended and actual function. Guided by a resource-aware mathematical model, we identify and engineer natural and synthetic miRNA-based incoherent feedforward loop (iFFL) circuits that mitigate gene expression burden. The implementation of these circuits features the use of endogenous miRNAs as elementary components of the engineered iFFL device, a versatile hybrid design that allows burden mitigation to be achieved across different cell-lines with minimal resource requirements. This study establishes the foundations for context-aware prediction and improvement of in vivo synthetic circuit performance, paving the way towards more rational synthetic construct design in mammalian cells.


Assuntos
Expressão Gênica , Mamíferos/genética , MicroRNAs/genética , Animais , Redes Reguladoras de Genes , Humanos , Mamíferos/metabolismo , Proteínas/genética , Proteínas/metabolismo
6.
Nucleic Acids Res ; 48(15): 8724-8739, 2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32735645

RESUMO

T cell activation is a well-established model for studying cellular responses to exogenous stimulation. Motivated by our previous finding that intron retention (IR) could lead to transcript instability, in this study, we performed BruChase-Seq to experimentally monitor the expression dynamics of nascent transcripts in resting and activated CD4+ T cells. Computational modeling was then applied to quantify the stability of spliced and intron-retained transcripts on a genome-wide scale. Beyond substantiating that intron-retained transcripts were considerably less stable than spliced transcripts, we found a global stabilization of spliced mRNAs upon T cell activation, although the stability of intron-retained transcripts remained relatively constant. In addition, we identified that La-related protein 4 (LARP4), an RNA-binding protein (RBP) known to enhance mRNA stability, was involved in T cell activation-dependent mRNA stabilization. Knocking out Larp4 in mice destabilized Nfκb1 mRNAs and reduced secretion of interleukin-2 (IL2) and interferon-gamma (IFNγ), two factors critical for T cell proliferation and function. We propose that coordination between splicing regulation and mRNA stability may provide a novel paradigm to control spatiotemporal gene expression during T cell activation.


Assuntos
Interferon gama/genética , Interleucina-2/genética , Proteínas/genética , Estabilidade de RNA/genética , Transcriptoma/genética , Processamento Alternativo/genética , Animais , Humanos , Íntrons/genética , Ativação Linfocitária/genética , Camundongos , NF-kappa B/genética , Ligação Proteica/genética , RNA Mensageiro/genética , Linfócitos T/metabolismo
7.
Nat Commun ; 11(1): 4171, 2020 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-32820176

RESUMO

Spiralia is a large, ancient and diverse clade of animals, with a conserved early developmental program but diverse larval and adult morphologies. One trait shared by many spiralians is the presence of ciliary bands used for locomotion and feeding. To learn more about spiralian-specific traits we have examined the expression of 20 genes with protein motifs that are strongly conserved within the Spiralia, but not detectable outside of it. Here, we show that two of these are specifically expressed in the main ciliary band of the mollusc Tritia (also known as Ilyanassa). Their expression patterns in representative species from five more spiralian phyla-the annelids, nemerteans, phoronids, brachiopods and rotifers-show that at least one of these, lophotrochin, has a conserved and specific role in particular ciliated structures, most consistently in ciliary bands. These results highlight the potential importance of lineage-specific genes or protein motifs for understanding traits shared across ancient lineages.


Assuntos
Motivos de Aminoácidos/genética , Cílios/genética , Invertebrados/genética , Proteínas/genética , Animais , Anelídeos/classificação , Anelídeos/genética , Anelídeos/fisiologia , Evolução Biológica , Cílios/fisiologia , Comportamento Alimentar/fisiologia , Perfilação da Expressão Gênica/métodos , Invertebrados/classificação , Invertebrados/fisiologia , Larva/genética , Larva/fisiologia , Locomoção/fisiologia , Moluscos/classificação , Moluscos/genética , Moluscos/fisiologia , Filogenia , Especificidade da Espécie
8.
PLoS Genet ; 16(8): e1008691, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32764743

RESUMO

Primary ciliary dyskinesia (PCD) is characterized by chronic airway disease, reduced fertility, and randomization of the left/right body axis. It is caused by defects of motile cilia and sperm flagella. We screened a cohort of affected individuals that lack an obvious axonemal defect for pathogenic variants using whole exome capture, next generation sequencing, and bioinformatic analysis assuming an autosomal recessive trait. We identified one subject with an apparently homozygous nonsense variant [(c.1762C>T), p.(Arg588*)] in the uncharacterized CFAP57 gene. Interestingly, the variant results in the skipping of exon 11 (58 amino acids), which may be due to disruption of an exonic splicing enhancer. In normal human nasal epithelial cells, CFAP57 localizes throughout the ciliary axoneme. Nasal cells from the PCD patient express a shorter, mutant version of CFAP57 and the protein is not incorporated into the axoneme. The missing 58 amino acids include portions of WD repeats that may be important for loading onto the intraflagellar transport (IFT) complexes for transport or docking onto the axoneme. A reduced beat frequency and an alteration in ciliary waveform was observed. Knockdown of CFAP57 in human tracheobronchial epithelial cells (hTECs) recapitulates these findings. Phylogenetic analysis showed that CFAP57 is highly conserved in organisms that assemble motile cilia. CFAP57 is allelic with the BOP2/IDA8/FAP57 gene identified previously in Chlamydomonas reinhardtii. Two independent, insertional fap57 Chlamydomonas mutant strains show reduced swimming velocity and altered waveforms. Tandem mass tag (TMT) mass spectroscopy shows that FAP57 is missing, and the "g" inner dyneins (DHC7 and DHC3) and the "d" inner dynein (DHC2) are reduced, but the FAP57 paralog FBB7 is increased. Together, our data identify a homozygous variant in CFAP57 that causes PCD that is likely due to a defect in the inner dynein arm assembly process.


Assuntos
Axonema/metabolismo , Transtornos da Motilidade Ciliar/genética , Códon sem Sentido , Dineínas/metabolismo , Proteínas/genética , Células 3T3 , Adulto , Animais , Axonema/fisiologia , Células Cultivadas , Chlamydomonas reinhardtii , Cílios/metabolismo , Cílios/fisiologia , Transtornos da Motilidade Ciliar/patologia , Sequência Conservada , Humanos , Masculino , Camundongos , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas/química , Proteínas/metabolismo , Mucosa Respiratória/metabolismo
9.
PLoS One ; 15(8): e0233247, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32857759

RESUMO

Poly(glycine-alanine) (polyGA) is one of the polydipeptides expressed in Frontotemporal Dementia and/or Amyotrophic Lateral Sclerosis 1 caused by C9ORF72 mutations and accumulates as inclusion bodies in the brain of patients. Superficially these inclusions are similar to those formed by polyglutamine (polyQ)-expanded Huntingtin exon 1 (Httex1) in Huntington's disease. Both have been reported to form an amyloid-like structure suggesting they might aggregate via similar mechanisms and therefore recruit the same repertoire of endogenous proteins. When co-expressed in the same cell, polyGA101 and Httex1(Q97) inclusions adopted immiscible phases suggesting different endogenous proteins would be enriched. Proteomic analyses identified 822 proteins in the inclusions. Only 7 were specific to polyGA and 4 specific to Httex1(Q97). Quantitation demonstrated distinct enrichment patterns for the proteins not specific to each inclusion type (up to ~8-fold normalized to total mass). The proteasome, microtubules, TriC chaperones, and translational machinery were enriched in polyGA aggregates, whereas Dnaj chaperones, nuclear envelope and RNA splicing proteins were enriched in Httex1(Q97) aggregates. Both structures revealed a collection of folding and degradation machinery including proteins in the Httex1(Q97) aggregates that are risk factors for other neurodegenerative diseases involving protein aggregation when mutated, which suggests a convergence point in the pathomechanisms of these diseases.


Assuntos
Corpos de Inclusão/metabolismo , Peptídeos/metabolismo , Proteínas/metabolismo , Animais , Proteína C9orf72/genética , Proteína C9orf72/metabolismo , Linhagem Celular , Éxons , Humanos , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Corpos de Inclusão/genética , Corpos de Inclusão/patologia , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Camundongos , Microscopia Confocal , Mutação , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Peptídeos/genética , Agregação Patológica de Proteínas/genética , Agregação Patológica de Proteínas/metabolismo , Agregação Patológica de Proteínas/patologia , Proteínas/genética , Proteólise , Proteoma/genética , Proteoma/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Fatores de Risco , Solubilidade
10.
PLoS One ; 15(8): e0237814, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32804975

RESUMO

Schaaf-Yang syndrome (SYS) is a neurodevelopmental disorder caused by truncating variants in the paternal allele of MAGEL2, located in the Prader-Willi critical region, 15q11-q13. Although the phenotypes of SYS overlap those of Prader-Willi syndrome (PWS), including neonatal hypotonia, feeding problems, and developmental delay/intellectual disability, SYS patients show autism spectrum disorder and joint contractures, which are atypical phenotypes for PWS. Therefore, we hypothesized that the truncated Magel2 protein could potentially produce gain-of-function toxic effects. To test the hypothesis, we generated two engineered mouse models; one, an overexpression model that expressed the N-terminal region of Magel2 that was FLAG tagged with a strong ubiquitous promoter, and another, a genome-edited model that carried a truncating variant in Magel2 generated using the CRISPR/Cas9 system. In the overexpression model, all transgenic mice died in the fetal or neonatal period indicating embryonic or neonatal lethality of the transgene. Therefore, overexpression of the truncated Magel2 could show toxic effects. In the genome-edited model, we generated a mouse model carrying a frameshift variant (c.1690_1924del; p(Glu564Serfs*130)) in Magel2. Model mice carrying the frameshift variant in the paternal or maternal allele of Magel2 were termed Magel2P:fs and Magel2M:fs, respectively. The imprinted expression and spatial distribution of truncating Magel2 transcripts in the brain were maintained. Although neonatal Magel2P:fs mice were lighter than wildtype littermates, Magel2P:fs males and females weighed the same as their wildtype littermates by eight and four weeks of age, respectively. Collectively, the overexpression mouse model may recapitulate fetal or neonatal death, which are the severest phenotypes for SYS. In contrast, the genome-edited mouse model maintains genomic imprinting and distribution of truncated Magel2 transcripts in the brain, but only partially recapitulates SYS phenotypes. Therefore, our results imply that simple gain-of-function toxic effects may not explain the patho-mechanism of SYS, but rather suggest a range of effects due to Magel2 variants as in human SYS patients.


Assuntos
Antígenos de Neoplasias/genética , Mutação/genética , Proteínas/genética , Animais , Antígenos de Neoplasias/química , Antígenos de Neoplasias/metabolismo , Peso Corporal , Encéfalo/metabolismo , Modelos Animais de Doenças , Feminino , Edição de Genes , Regulação da Expressão Gênica , Células HEK293 , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Linhagem , Fenótipo , Proteínas/química , Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
11.
Nucleic Acids Res ; 48(15): 8302-8319, 2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32725210

RESUMO

We used the high resolution and accuracy of the Cambridge Structural Database (CSD) to provide detailed information regarding base pairing interactions of selected nucleobases. We searched for base pairs in which nucleobases interact with each other through two or more hydrogen bonds and form more or less planar structures. The investigated compounds were either free forms or derivatives of adenine, guanine, hypoxanthine, thymine, uracil and cytosine. We divided our findings into categories including types of pairs, protonation patterns and whether they are formed by free bases or substituted ones. We found base pair types that are exclusive to small molecule crystal structures, some that can be found only in RNA containing crystal structures and many that are native to both environments. With a few exceptions, nucleobase protonation generally followed a standard pattern governed by pKa values. The lengths of hydrogen bonds did not depend on whether the nucleobases forming a base pair were charged or not. The reasons why particular nucleobases formed base pairs in a certain way varied significantly.


Assuntos
Bases de Dados de Proteínas , Ligação de Hidrogênio , Conformação Proteica , Proteínas/genética , Adenina/química , Pareamento de Bases/genética , Cristalografia por Raios X , Citosina/química , Guanina/química , Hipoxantina/química , Estrutura Molecular , Proteínas/química , Proteínas/ultraestrutura , Bibliotecas de Moléculas Pequenas/química , Timina/química , Uracila/química
12.
Nat Commun ; 11(1): 3606, 2020 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-32681016

RESUMO

Mitochondrial metabolism has emerged as a promising target against the mechanisms of tumor growth. Herein, we have screened an FDA-approved library to identify drugs that inhibit mitochondrial respiration. The ß1-blocker nebivolol specifically hinders oxidative phosphorylation in cancer cells by concertedly inhibiting Complex I and ATP synthase activities. Complex I inhibition is mediated by interfering the phosphorylation of NDUFS7. Inhibition of the ATP synthase is exerted by the overexpression and binding of the ATPase Inhibitory Factor 1 (IF1) to the enzyme. Remarkably, nebivolol also arrests tumor angiogenesis by arresting endothelial cell proliferation. Altogether, targeting mitochondria and angiogenesis triggers a metabolic and oxidative stress crisis that restricts the growth of colon and breast carcinomas. Nebivolol holds great promise to be repurposed for the treatment of cancer patients.


Assuntos
Antagonistas Adrenérgicos/farmacologia , Indutores da Angiogênese/farmacologia , Neoplasias da Mama/fisiopatologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/fisiopatologia , Mitocôndrias/efeitos dos fármacos , Nebivolol/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/genética , Feminino , Humanos , Masculino , Mitocôndrias/genética , Mitocôndrias/metabolismo , ATPases Mitocondriais Próton-Translocadoras/genética , ATPases Mitocondriais Próton-Translocadoras/metabolismo , NADH Desidrogenase/genética , NADH Desidrogenase/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Proteínas/genética , Proteínas/metabolismo
13.
Cytogenet Genome Res ; 160(6): 295-308, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32683365

RESUMO

Intramolecular coevolution of amino acid sites has repeatedly been studied to improve predictions on protein structure and function. Thereby, the focus was on bacterial proteins with available crystallographic data. However, intramolecular coevolution has not yet been compared between protein sets along a gradient of functional proximity to fertilization. This is especially true for the potential effect of external selective forces on intraprotein coevolution. In this study, we investigated both aspects in equally sized sets of mammalian proteins representing spermatozoa, testis, entire body, and liver. For coevolutionary analyses, we derived the proportion of covarying sites per protein from amino acid alignments of 10 mammalian orthologues each. In confirmation of the validity of our coevolution proxy, we found positive associations with the nonsynonymous or amino acid substitution rate in all protein sets. However, our coevolution proxy negatively correlated with the number of protein interactants (node degree) in male reproductive protein sets alone. In addition, a negative association of our coevolution proxy with protein hydrophobicity was significant in sperm proteins only. Accordingly, the restrictive effect of protein interactants was most pronounced in male reproductive proteins, and the tendency of sperm proteins to form internal structures decreased the more coevolutionary sites they had. Both aspects illustrate that the share of outward and thus functional coevolution increases with greater proximity to fertilization. We found this conclusion confirmed by additional comparisons within sperm proteins. Thus, sperm proteins with high hydrophobicity had the lowest proportions of covarying sites and, according to gene annotations, localized more frequently to internal cellular structures. They should therefore be less exposed to postcopulatory forms of sexual selection. Their counterparts with low hydrophobicity had larger proportions of covarying sites and more often resided at the cell membrane or were secreted. At the cellular level, they are thus closer to externally induced forces of postcopulatory selection which are known for their potential to increase substitution rates. In addition, we show that the intermediary status of the testicular protein set in correlation analyses is probably due to a special combination of reproductive and somatic involvements.


Assuntos
Evolução Molecular , Fertilização , Proteínas/química , Proteínas/metabolismo , Espermatozoides/química , Espermatozoides/metabolismo , Animais , Doença , Fertilização/genética , Humanos , Interações Hidrofóbicas e Hidrofílicas , Masculino , Proteínas de Membrana/química , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Proteínas/genética , Proteoma/química , Proteoma/metabolismo , Suínos
14.
Arch Virol ; 165(10): 2279-2289, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32719955

RESUMO

In the early stage of virus infection, the pattern recognition receptor (PRR) signaling pathway of the host cell is activated to induce interferon production, activating interferon-stimulated genes (ISGs) that encode antiviral proteins that exert antiviral effects. Viperin is one of the innate antiviral proteins that exert broad-spectrum antiviral effects by various mechanisms. Porcine epidemic diarrhea virus (PEDV) is a coronavirus that causes huge losses to the pig industry. Research on early antiviral responses in the gastrointestinal tract is essential for developing strategies to prevent the spread of PEDV. In this study, we investigated the mechanisms of viperin in PEDV-infected IPEJ-C2 cells. Increased expression of interferon and viperin and decreased replication of PEDV with a clear reduction in the viral load were observed in PEDV-infected IPEC-J2 cells. Amino acids 1-50 of porcine viperin contain an endoplasmic reticulum signal sequence that allows viperin to be anchored to the endoplasmic reticulum and are necessary for its function in inhibiting PEDV proliferation. The interaction of the viperin S-adenosylmethionine domain with the N protein of PEDV was confirmed via confocal laser scanning microscopy and co-immunoprecipitation. This interaction might interfere with viral replication or assembly to reduce virus proliferation. Our results highlight a potential mechanism whereby viperin is able to inhibit PEDV replication and play an antiviral role in innate immunity.


Assuntos
Antivirais/metabolismo , Interações entre Hospedeiro e Microrganismos/fisiologia , Proteínas do Nucleocapsídeo/fisiologia , Vírus da Diarreia Epidêmica Suína/fisiologia , Animais , Linhagem Celular , Infecções por Coronavirus/genética , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Interações entre Hospedeiro e Microrganismos/genética , Interações entre Hospedeiro e Microrganismos/imunologia , Imunidade Inata , Interferons/biossíntese , Camundongos , Camundongos Endogâmicos BALB C , Proteínas do Nucleocapsídeo/antagonistas & inibidores , Proteínas do Nucleocapsídeo/química , Vírus da Diarreia Epidêmica Suína/imunologia , Vírus da Diarreia Epidêmica Suína/patogenicidade , Domínios e Motivos de Interação entre Proteínas , Proteínas/química , Proteínas/genética , Proteínas/fisiologia , Interferência de RNA , Suínos , Replicação Viral
15.
Am J Hum Genet ; 107(2): 342-351, 2020 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-32673564

RESUMO

Male infertility affects ∼7% of men, but its causes remain poorly understood. The most severe form is non-obstructive azoospermia (NOA), which is, in part, caused by an arrest at meiosis. So far, only a few validated disease-associated genes have been reported. To address this gap, we performed whole-exome sequencing in 58 men with unexplained meiotic arrest and identified the same homozygous frameshift variant c.676dup (p.Trp226LeufsTer4) in M1AP, encoding meiosis 1 associated protein, in three unrelated men. This variant most likely results in a truncated protein as shown in vitro by heterologous expression of mutant M1AP. Next, we screened four large cohorts of infertile men and identified three additional individuals carrying homozygous c.676dup and three carrying combinations of this and other likely causal variants in M1AP. Moreover, a homozygous missense variant, c.1166C>T (p.Pro389Leu), segregated with infertility in five men from a consanguineous Turkish family. The common phenotype between all affected men was NOA, but occasionally spermatids and rarely a few spermatozoa in the semen were observed. A similar phenotype has been described for mice with disruption of M1ap. Collectively, these findings demonstrate that mutations in M1AP are a relatively frequent cause of autosomal recessive severe spermatogenic failure and male infertility with strong clinical validity.


Assuntos
Pontos de Checagem do Ciclo Celular/genética , Infertilidade Masculina/genética , Meiose/genética , Mutação/genética , Proteínas/genética , Espermatogênese/genética , Adulto , Alelos , Animais , Azoospermia/genética , Homozigoto , Humanos , Masculino , Camundongos , Fenótipo , Espermatozoides/anormalidades , Testículo/anormalidades , Turquia , Sequenciamento Completo do Exoma/métodos
16.
Nature ; 584(7819): 142-147, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32612238

RESUMO

Nuclear processes, such as V(D)J recombination, are orchestrated by the three-dimensional organization of chromosomes at multiple levels, including compartments1 and topologically associated domains (TADs)2,3 consisting of chromatin loops4. TADs are formed by chromatin-loop extrusion5-7, which depends on the loop-extrusion function of the ring-shaped cohesin complex8-12. Conversely, the cohesin-release factor Wapl13,14 restricts loop extension10,15. The generation of a diverse antibody repertoire, providing humoral immunity to pathogens, requires the participation of all V genes in V(D)J recombination16, which depends on contraction of the 2.8-Mb-long immunoglobulin heavy chain (Igh) locus by Pax517,18. However, how Pax5 controls Igh contraction in pro-B cells remains unknown. Here we demonstrate that locus contraction is caused by loop extrusion across the entire Igh locus. Notably, the expression of Wapl is repressed by Pax5 specifically in pro-B and pre-B cells, facilitating extended loop extrusion by increasing the residence time of cohesin on chromatin. Pax5 mediates the transcriptional repression of Wapl through a single Pax5-binding site by recruiting the polycomb repressive complex 2 to induce bivalent chromatin at the Wapl promoter. Reduced Wapl expression causes global alterations in the chromosome architecture, indicating that the potential to recombine all V genes entails structural changes of the entire genome in pro-B cells.


Assuntos
Genes de Cadeia Pesada de Imunoglobulina/genética , Cadeias Pesadas de Imunoglobulinas/genética , Região Variável de Imunoglobulina/genética , Fator de Transcrição PAX5/metabolismo , Proteínas/genética , Proteínas Repressoras/metabolismo , Recombinação V(D)J/genética , Animais , Linfócitos B/citologia , Linfócitos B/metabolismo , Sítios de Ligação , Proteínas de Ciclo Celular/metabolismo , Montagem e Desmontagem da Cromatina , Proteínas Cromossômicas não Histona/metabolismo , Cadeias Pesadas de Imunoglobulinas/química , Região Variável de Imunoglobulina/química , Camundongos , Complexo Repressor Polycomb 2/metabolismo , Células Precursoras de Linfócitos B/citologia , Células Precursoras de Linfócitos B/metabolismo , Regiões Promotoras Genéticas/genética
17.
BMC Bioinformatics ; 21(1): 275, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32611389

RESUMO

BACKGROUND: Protein engineering has many applications for industry, such as the development of new drugs, vaccines, treatment therapies, food, and biofuel production. A common way to engineer a protein is to perform mutations in functionally essential residues to optimize their function. However, the discovery of beneficial mutations for proteins is a complex task, with a time-consuming and high cost for experimental validation. Hence, computational approaches have been used to propose new insights for experiments narrowing the search space and reducing the costs. RESULTS: In this study, we developed Proteus (an acronym for Protein Engineering Supporter), a new algorithm for proposing mutation pairs in a target 3D structure. These suggestions are based on contacts observed in other known structures from Protein Data Bank (PDB). Proteus' basic assumption is that if a non-interacting pair of amino acid residues in the target structure is exchanged to an interacting pair, this could enhance protein stability. This trade is only allowed if the main-chain conformation of the residues involved in the contact is conserved. Furthermore, no steric impediment is expected between the proposed mutations and the surrounding protein atoms. To evaluate Proteus, we performed two case studies with proteins of industrial interests. In the first case study, we evaluated if the mutations suggested by Proteus for four protein structures enhance the number of inter-residue contacts. Our results suggest that most mutations proposed by Proteus increase the number of interactions into the protein. In the second case study, we used Proteus to suggest mutations for a lysozyme protein. Then, we compared Proteus' outcomes to mutations with available experimental evidence reported in the ProTherm database. Four mutations, in which our results agree with the experimental data, were found. This could be initial evidence that changes in the side-chain of some residues do not cause disturbances that harm protein structure stability. CONCLUSION: We believe that Proteus could be used combined with other methods to give new insights into the rational development of engineered proteins. Proteus user-friendly web-based tool is available at < http://proteus.dcc.ufmg.br >.


Assuntos
Proteínas/química , Interface Usuário-Computador , Algoritmos , Bases de Dados de Proteínas , Muramidase/química , Muramidase/genética , Muramidase/metabolismo , Mutagênese , Engenharia de Proteínas/métodos , Estrutura Terciária de Proteína , Proteínas/genética , Proteínas/metabolismo
18.
Nat Commun ; 11(1): 3368, 2020 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-32632093

RESUMO

Blood pressure (BP) was inconsistently associated with migraine and the mechanisms of BP-lowering medications in migraine prophylaxis are unknown. Leveraging large-scale summary statistics for migraine (Ncases/Ncontrols = 59,674/316,078) and BP (N = 757,601), we find positive genetic correlations of migraine with diastolic BP (DBP, rg = 0.11, P = 3.56 × 10-06) and systolic BP (SBP, rg = 0.06, P = 0.01), but not pulse pressure (PP, rg = -0.01, P = 0.75). Cross-trait meta-analysis reveals 14 shared loci (P ≤ 5 × 10-08), nine of which replicate (P < 0.05) in the UK Biobank. Five shared loci (ITGB5, SMG6, ADRA2B, ANKDD1B, and KIAA0040) are reinforced in gene-level analysis and highlight potential mechanisms involving vascular development, endothelial function and calcium homeostasis. Mendelian randomization reveals stronger instrumental estimates of DBP (OR [95% CI] = 1.20 [1.15-1.25]/10 mmHg; P = 5.57 × 10-25) on migraine than SBP (1.05 [1.03-1.07]/10 mmHg; P = 2.60 × 10-07) and a corresponding opposite effect for PP (0.92 [0.88-0.95]/10 mmHg; P = 3.65 × 10-07). These findings support a critical role of DBP in migraine susceptibility and shared biology underlying BP and migraine.


Assuntos
Pressão Sanguínea/genética , Predisposição Genética para Doença/genética , Estudo de Associação Genômica Ampla/métodos , Metanálise como Assunto , Transtornos de Enxaqueca/genética , Polimorfismo de Nucleotídeo Único , Humanos , Hipertensão/genética , Cadeias beta de Integrinas/genética , Análise da Randomização Mendeliana/métodos , Proteínas/genética , Receptores Adrenérgicos alfa 2/genética , Fatores de Risco , Telomerase/genética
19.
Nat Commun ; 11(1): 3409, 2020 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-32641778

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) is associated with high mortality and therapy resistance. Here, we show that low expression of κB-Ras GTPases is frequently detected in PDAC and correlates with higher histologic grade. In a model of KRasG12D-driven PDAC, loss of κB-Ras accelerates tumour development and shortens median survival. κB-Ras deficiency promotes acinar-to-ductal metaplasia (ADM) during tumour initiation as well as tumour progression through intrinsic effects on proliferation and invasion. κB-Ras proteins are also required for acinar regeneration after pancreatitis, demonstrating a general role in control of plasticity. Molecularly, upregulation of Ral GTPase activity and Sox9 expression underlies the observed phenotypes, identifying a previously unrecognized function of Ral signalling in ADM. Our results provide evidence for a tumour suppressive role of κB-Ras proteins and highlight low κB-Ras levels and consequent loss of Ral control as risk factors, thus emphasizing the necessity for therapeutic options that allow interference with Ral-driven signalling.


Assuntos
Células Acinares/metabolismo , Carcinoma Ductal Pancreático/genética , GTP Fosfo-Hidrolases/genética , Neoplasias Pancreáticas/genética , Pancreatite/genética , Proteínas/genética , Células Acinares/patologia , Idoso , Animais , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Feminino , GTP Fosfo-Hidrolases/metabolismo , Regulação da Expressão Gênica , Humanos , Proteínas I-kappa B/genética , Proteínas I-kappa B/metabolismo , Estimativa de Kaplan-Meier , Masculino , Metaplasia/genética , Metaplasia/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Pancreatite/metabolismo , Proteínas/metabolismo , Fatores de Transcrição SOX9/genética , Fatores de Transcrição SOX9/metabolismo , Proteínas ral de Ligação ao GTP/genética , Proteínas ral de Ligação ao GTP/metabolismo , Proteínas ras/genética , Proteínas ras/metabolismo
20.
PLoS One ; 15(7): e0235706, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32639988

RESUMO

During type 1 immune responses, CD4 T helper 1 (Th1) cells and CD8 T cells are activated via IL-12 and contribute to the elimination of intracellular pathogens through interferon gamma (IFNγ) production. In this study, we identified Placenta-specific 8 (Plac8) as a gene that is uniquely expressed in Th1 CD4 T cells relative to other CD4 T cell subsets and hypothesized that Plac8 may represent a novel therapeutic target in Th1 CD4 T cells. First, we determined that Plac8 mRNA in CD4 T cells was induced following IL-12 stimulation via an indirect route that required new protein synthesis. Upon evaluating the functional relevance of Plac8 expression in Th1 CD4 T cells, we discovered that Plac8 was important for suppressing IFNγ mRNA and protein production by CD4 T cells 24 hours after IL-12 stimulation, however Plac8 did not contribute to pathogenic CD4 T cell function during two models of intestinal inflammation. We also noted relatively high basal expression of Plac8 in CD8 T cells which could be further induced following IL-12 stimulation in CD8 T cells. Furthermore, Plac8 expression was important for establishing an optimal CD8 T cell response against influenza A virus via a T cell-intrinsic manner. Altogether, these results implicate Plac8 as a potential regulator of Th1 CD4 and CD8 T cell responses during Th1 T cell-driven inflammation.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Interferon gama/metabolismo , Proteínas/metabolismo , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Colite/imunologia , Colite/patologia , Modelos Animais de Doenças , Interferon gama/genética , Interleucina-12/farmacologia , Antígenos Comuns de Leucócito/genética , Antígenos Comuns de Leucócito/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/patologia , Proteínas/genética , RNA Mensageiro/metabolismo , Regulação para Cima/efeitos dos fármacos
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