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1.
Nat Commun ; 11(1): 5536, 2020 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-33139700

RESUMO

MAVS and MITA are essential adaptor proteins mediating innate antiviral immune responses against RNA and DNA viruses, respectively. Here we show that RNF115 plays dual roles in response to RNA or DNA virus infections by catalyzing distinct types of ubiquitination of MAVS and MITA at different phases of viral infection. RNF115 constitutively interacts with and induces K48-linked ubiquitination and proteasomal degradation of homeostatic MAVS in uninfected cells, whereas associates with and catalyzes K63-linked ubiquitination of MITA after HSV-1 infection. Consistently, the protein levels of MAVS are substantially increased in Rnf115-/- organs or cells without viral infection, and HSV-1-induced aggregation of MITA is impaired in Rnf115-/- cells compared to the wild-type counterparts. Consequently, the Rnf115-/- mice exhibit hypo- and hyper-sensitivity to EMCV and HSV-1 infection, respectively. These findings highlight dual regulation of cellular antiviral responses by RNF115-mediated ubiquitination of MAVS and MITA and contribute to our understanding of innate immune signaling.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Infecções por Cardiovirus/imunologia , Herpes Simples/imunologia , Imunidade Inata , Proteínas de Membrana/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Infecções por Cardiovirus/patologia , Infecções por Cardiovirus/virologia , Modelos Animais de Doenças , Vírus da Encefalomiocardite/imunologia , Feminino , Células HEK293 , Herpes Simples/patologia , Herpes Simples/virologia , Herpesvirus Humano 1/imunologia , Interações Hospedeiro-Patógeno/imunologia , Humanos , Lisina/metabolismo , Macrófagos/imunologia , Macrófagos/virologia , Masculino , Camundongos , Camundongos Knockout , Cultura Primária de Células , Agregados Proteicos/imunologia , RNA Interferente Pequeno/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/fisiologia , Ubiquitinação/imunologia
2.
Zhongguo Xue Xi Chong Bing Fang Zhi Za Zhi ; 32(5): 542-547, 2020 May 08.
Artigo em Chinês | MEDLINE | ID: mdl-33185072

RESUMO

Aquaporins (AQPs) are a class of membrane intrinsic proteins in medical helminthes that specifically mediate the transmembrane transport of water or other solute molecules. Previous studies have demonstrated that AQPs play a critical role in promoting the transmembrane transport of water, osmoregulation, uptake of nutrients, release of toxic metabolic products and transport of antiparasitic drugs, which may serve as promising vaccine candidates and drug targets for parasitic diseases. This review describes the structural characteristics of AQPs in medical helminthes, and discusses the feasibility of these AQPs as antihelminth vaccine candidates and drug targets, so as to provide insights into the development of novel vaccines and drugs against parasitic diseases.


Assuntos
Aquaporinas , Helmintíase , Helmintos , Animais , Aquaporinas/metabolismo , Transporte Biológico , Helmintíase/parasitologia , Helmintíase/prevenção & controle , Helmintos/fisiologia , Humanos , Proteínas de Membrana/metabolismo , Pesquisa/tendências , Água/metabolismo
3.
BMC Bioinformatics ; 21(1): 503, 2020 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-33153432

RESUMO

BACKGROUND: The formation of contacts among protein secondary structure elements (SSEs) is an important step in protein folding as it determines topology of protein tertiary structure; hence, inferring inter-SSE contacts is crucial to protein structure prediction. One of the existing strategies infers inter-SSE contacts directly from the predicted possibilities of inter-residue contacts without any preprocessing, and thus suffers from the excessive noises existing in the predicted inter-residue contacts. Another strategy defines SSEs based on protein secondary structure prediction first, and then judges whether each candidate SSE pair could form contact or not. However, it is difficult to accurately determine boundary of SSEs due to the errors in secondary structure prediction. The incorrectly-deduced SSEs definitely hinder subsequent prediction of the contacts among them. RESULTS: We here report an accurate approach to infer the inter-SSE contacts (thus called as ISSEC) using the deep object detection technique. The design of ISSEC is based on the observation that, in the inter-residue contact map, the contacting SSEs usually form rectangle regions with characteristic patterns. Therefore, ISSEC infers inter-SSE contacts through detecting such rectangle regions. Unlike the existing approach directly using the predicted probabilities of inter-residue contact, ISSEC applies the deep convolution technique to extract high-level features from the inter-residue contacts. More importantly, ISSEC does not rely on the pre-defined SSEs. Instead, ISSEC enumerates multiple candidate rectangle regions in the predicted inter-residue contact map, and for each region, ISSEC calculates a confidence score to measure whether it has characteristic patterns or not. ISSEC employs greedy strategy to select non-overlapping regions with high confidence score, and finally infers inter-SSE contacts according to these regions. CONCLUSIONS: Comprehensive experimental results suggested that ISSEC outperformed the state-of-the-art approaches in predicting inter-SSE contacts. We further demonstrated the successful applications of ISSEC to improve prediction of both inter-residue contacts and tertiary structure as well.


Assuntos
Algoritmos , Proteínas/química , Bases de Dados de Proteínas , Proteínas de Membrana/química , Conformação Proteica em Folha beta , Estrutura Secundária de Proteína
4.
Nat Commun ; 11(1): 5436, 2020 10 28.
Artigo em Inglês | MEDLINE | ID: mdl-33116123

RESUMO

Harmful effects of high fructose intake on health have been widely reported. Although fructose is known to promote cancer, little is known about the underlying mechanisms. Here, we found that fructose triggers breast cancer metastasis through the ketohexokinase-A signaling pathway. Molecular experiments showed that ketohexokinase-A, rather than ketohexokinase-C, is necessary and sufficient for fructose-induced cell invasion. Ketohexokinase-A-overexpressing breast cancer was found to be highly metastatic in fructose-fed mice. Mechanistically, cytoplasmic ketohexokinase-A enters into the nucleus during fructose stimulation, which is mediated by LRRC59 and KPNB1. In the nucleus, ketohexokinase-A phosphorylates YWHAH at Ser25 and the YWHAH recruits SLUG to the CDH1 promoter, which triggers cell migration. This study provides the effect of nutrition on breast cancer metastasis. High intake of fructose should be restricted in cancer patients to reduce the risk of metastasis. From a therapeutic perspective, the ketohexokinase-A signaling pathway could be a potential target to prevent cancer metastasis.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Frutoquinases/metabolismo , Frutose/administração & dosagem , Frutose/metabolismo , Proteínas 14-3-3/antagonistas & inibidores , Proteínas 14-3-3/genética , Proteínas 14-3-3/metabolismo , Animais , Carcinógenos/administração & dosagem , Carcinógenos/metabolismo , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Feminino , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Mamárias Experimentais/metabolismo , Neoplasias Mamárias Experimentais/patologia , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Invasividade Neoplásica/patologia , Metástase Neoplásica/patologia , Fosforilação , Transdução de Sinais , beta Carioferinas/metabolismo
5.
Medicine (Baltimore) ; 99(43): e22000, 2020 Oct 23.
Artigo em Inglês | MEDLINE | ID: mdl-33120728

RESUMO

The expression of tumor stem cell markers musashi1 (msi1) and numb in brain metastases were detected to explore their roles in the development of brain metastases.A total of 51 cases of brain metastasis, 29 cases of primary tumor and 15 cases of normal brain tissue were selected. Immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR) were used to detect msi1 and numb expression at the protein and mRNA levels. Correlation between msi1 and numb in brain metastases were evaluated.Immunohistochemistry and RT-PCR showed that no significant difference in the expression of msi1 and numb between brain metastases and primary tumors was observed (P > .05); the expression of msi1 and numb in brain metastases was significantly higher than that in normal brain tissues (P < .05); and the expression of msi1 and numb in primary tumors was significantly higher than that in normal brain tissues (P < .05). In general, the expression of msi1 gene was negatively correlated with the expression of numb at mRNA level by Pearson correlation analysis (r = -0.345, P < .05). Additionally, the expression of msi1 and numb in brain metastases was not related to gender, age, and tissue origin (P > .05).Msi1 is highly expressed in brain metastases and primary tumors, while numb is lowly expressed in brain metastases and primary tumors; msi1 and numb are negatively correlated in brain metastases, suggesting that msi1 and numb may have regulatory mechanisms in the development of brain metastases.


Assuntos
Neoplasias Encefálicas/genética , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/genética , Adulto , Idoso , Encéfalo/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/secundário , Estudos de Casos e Controles , Feminino , Humanos , Imuno-Histoquímica , Masculino , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/metabolismo , Proteínas de Ligação a RNA/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Adulto Jovem
6.
Nat Commun ; 11(1): 5459, 2020 10 29.
Artigo em Inglês | MEDLINE | ID: mdl-33122649

RESUMO

Biocontainment systems are needed to neutralize genetically modified organisms (GMOs) that pose ecological threats outside of controlled environments. In contrast, benign selection markers complement GMOs with reduced fitness. Benign selection agents serve as alternatives to antibiotics, which are costly and risk spread of antibiotic resistance. Here, we present a yeast biocontainment strategy leveraging engineered fluoride sensitivity and DNA vectors enabling use of fluoride as a selection agent. The biocontainment system addresses the scarcity of platforms available for yeast despite their prevalent use in industry and academia. In the absence of fluoride, the biocontainment strain exhibits phenotypes nearly identical to those of the wildtype strain. Low fluoride concentrations severely inhibit biocontainment strain growth, which is restored upon introduction of fluoride-based vectors. The biocontainment strategy is stringent, easily implemented, and applicable to several eukaryotes. Further, the DNA vectors enable genetic engineering at reduced costs and eliminate risks of propagating antibiotic resistance.


Assuntos
Fluoretos , Proteínas de Membrana/genética , Organismos Geneticamente Modificados/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Seleção Genética , Fluoretos/metabolismo , Fluoretos/farmacologia , Engenharia Genética , Marcadores Genéticos , Saccharomyces cerevisiae/efeitos dos fármacos , Biologia Sintética/métodos
7.
Nat Commun ; 11(1): 4990, 2020 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-33020478

RESUMO

Neurons are highly compartmentalized cells with tightly controlled subcellular protein organization. While brain transcriptome, connectome and global proteome maps are being generated, system-wide analysis of temporal protein dynamics at the subcellular level are currently lacking. Here, we perform a temporally-resolved surfaceome analysis of primary neuron cultures and reveal dynamic surface protein clusters that reflect the functional requirements during distinct stages of neuronal development. Direct comparison of surface and total protein pools during development and homeostatic synaptic scaling demonstrates system-wide proteostasis-independent remodeling of the neuronal surface, illustrating widespread regulation on the level of surface trafficking. Finally, quantitative analysis of the neuronal surface during chemical long-term potentiation (cLTP) reveals fast externalization of diverse classes of surface proteins beyond the AMPA receptor, providing avenues to investigate the requirement of exocytosis for LTP. Our resource (neurosurfaceome.ethz.ch) highlights the importance of subcellular resolution for systems-level understanding of cellular processes.


Assuntos
Proteínas de Membrana/metabolismo , Plasticidade Neuronal , Neurônios/metabolismo , Sinapses/metabolismo , Animais , Membrana Celular/metabolismo , Células Cultivadas , Potenciais Pós-Sinápticos Excitadores , Homeostase , Potenciação de Longa Duração , Mapas de Interação de Proteínas , Transporte Proteico , Proteostase , Ratos
8.
Nat Commun ; 11(1): 5140, 2020 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-33046712

RESUMO

Autism spectrum disorder (ASD) is thought to result from deviation from normal development of neural circuits and synaptic function. Many genes with mutation in ASD patients have been identified. Here we report that two molecules associated with ASD susceptibility, contactin associated protein-like 2 (CNTNAP2) and Abelson helper integration site-1 (AHI1), are required for synaptic function and ASD-related behavior in mice. Knockdown of CNTNAP2 or AHI1 in layer 2/3 pyramidal neurons of the developing mouse prefrontal cortex (PFC) reduced excitatory synaptic transmission, impaired social interaction and induced mild vocalization abnormality. Although the causes of reduced excitatory transmission were different, pharmacological enhancement of AMPA receptor function effectively restored impaired social behavior in both CNTNAP2- and AHI1-knockdown mice. We conclude that reduced excitatory synaptic transmission in layer 2/3 pyramidal neurons of the PFC leads to impaired social interaction and mild vocalization abnormality in mice.


Assuntos
Transtorno do Espectro Autista/metabolismo , Transtorno do Espectro Autista/psicologia , Córtex Pré-Frontal/metabolismo , Células Piramidais/metabolismo , Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Animais , Transtorno do Espectro Autista/genética , Transtorno do Espectro Autista/fisiopatologia , Comportamento Animal , Modelos Animais de Doenças , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Transmissão Sináptica
9.
Yonsei Med J ; 61(11): 923-934, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33107235

RESUMO

PURPOSE: The mechanisms of Wnt/ß-catenin pathway signaling and abnormal expression of tumor suppressor genes is not well known in gastric cancer (GC). Long non-coding RNA (lncRNA) has recently been identified as a possible link therein. In this study, we investigated the role of lung cancer associated transcript 1 (LUCAT1) in GC. MATERIALS AND METHODS: The expression of LUCAT1 in GC cell lines and 100 tissue samples was examined by qRT-PCR. Two different siRNAs were used for knockdown of LUCAT1 expression. Cell viability was assessed by MTT assay. To analyze metastasis, scratch wound-healing assay, a Matrigel invasion assay, and colony formation assay were performed. Apoptosis was analyzed by PI/Annexin-V staining. To check the methylation status in tumor suppressor genes, methylation-specific PCR was carried out. Western blot was performed to detect epithelial-mesenchymal transition and apoptosis markers upon silencing of LUCAT1 (siLUCAT1). RESULTS: LUCAT1 expression in GC cell lines and tissues was significantly elevated, compared to that in normal gastric cells and adjacent non-tumor tissues (p<0.001). Two different siRNAs for LUCAT1 reduced cell proliferation, invasion, and migration, compared to siCT (p<0.05), and these reductions were restored by pcDNA-LUCAT1 (p<0.05). siLUCAT1 elicited upregulation of the expression of CXXC4 and SFRP2. The expression of H3K27me3 was reduced by siLUCAT1, and this reduction was correlated with methylation of CXXC4 and SFRP2. Inhibition of LUCAT1 up-regulated EZH2 expression and resulted in demethylation of CXXC4 and SFRP2 through the Wnt/ß-catenin signaling pathway. CONCLUSION: We concluded that LUCAT1 induces methylation of CXXC4 and SFRP2, thereby regulating Wnt/ß-catenin signaling in GC.


Assuntos
Proteínas de Ligação a DNA/genética , Genes Supressores de Tumor , Proteínas de Membrana/genética , Invasividade Neoplásica/genética , RNA Longo não Codificante/metabolismo , Neoplasias Gástricas/genética , Fatores de Transcrição/genética , Apoptose/genética , Biomarcadores Tumorais/genética , Proliferação de Células/genética , Sobrevivência Celular , Proteínas de Ligação a DNA/metabolismo , Regulação para Baixo , Transição Epitelial-Mesenquimal , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Neoplasias Pulmonares/genética , Proteínas de Membrana/metabolismo , RNA Longo não Codificante/genética , RNA Interferente Pequeno/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Neoplasias Gástricas/patologia , Fatores de Transcrição/metabolismo , Regulação para Cima , Via de Sinalização Wnt
10.
Nat Commun ; 11(1): 5171, 2020 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-33057002

RESUMO

Excitatory and inhibitory neurons are connected into microcircuits that generate circuit output. Central in the hippocampal CA3 microcircuit is the mossy fiber (MF) synapse, which provides powerful direct excitatory input and indirect feedforward inhibition to CA3 pyramidal neurons. Here, we dissect its cell-surface protein (CSP) composition to discover novel regulators of MF synaptic connectivity. Proteomic profiling of isolated MF synaptosomes uncovers a rich CSP composition, including many CSPs without synaptic function and several that are uncharacterized. Cell-surface interactome screening identifies IgSF8 as a neuronal receptor enriched in the MF pathway. Presynaptic Igsf8 deletion impairs MF synaptic architecture and robustly decreases the density of bouton filopodia that provide feedforward inhibition. Consequently, IgSF8 loss impairs excitation/inhibition balance and increases excitability of CA3 pyramidal neurons. Our results provide insight into the CSP landscape and interactome of a specific excitatory synapse and reveal IgSF8 as a critical regulator of CA3 microcircuit connectivity and function.


Assuntos
Região CA3 Hipocampal/fisiologia , Proteínas de Transporte/metabolismo , Potenciais Pós-Sinápticos Excitadores/fisiologia , Proteínas de Membrana/metabolismo , Fibras Musgosas Hipocampais/metabolismo , Células Piramidais/fisiologia , Animais , Proteínas de Transporte/genética , Células Cultivadas , Células HEK293 , Humanos , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Técnicas de Patch-Clamp , Cultura Primária de Células , Proteômica , Ratos , Sinaptossomos/metabolismo
11.
Nat Commun ; 11(1): 5178, 2020 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-33057066

RESUMO

Japanese encephalitis virus (JEV) is a mosquito-borne zoonotic flavivirus that causes encephalitis and reproductive disorders in mammalian species. However, the host factors critical for its entry, replication, and assembly are poorly understood. Here, we design a porcine genome-scale CRISPR/Cas9 knockout (PigGeCKO) library containing 85,674 single guide RNAs targeting 17,743 protein-coding genes, 11,053 long ncRNAs, and 551 microRNAs. Subsequently, we use the PigGeCKO library to identify key host factors facilitating JEV infection in porcine cells. Several previously unreported genes required for JEV infection are highly enriched post-JEV selection. We conduct follow-up studies to verify the dependency of JEV on these genes, and identify functional contributions for six of the many candidate JEV-related host genes, including EMC3 and CALR. Additionally, we identify that four genes associated with heparan sulfate proteoglycans (HSPGs) metabolism, specifically those responsible for HSPGs sulfurylation, facilitate JEV entry into porcine cells. Thus, beyond our development of the largest CRISPR-based functional genomic screening platform for pig research to date, this study identifies multiple potentially vulnerable targets for the development of medical and breeding technologies to treat and prevent diseases caused by JEV.


Assuntos
Vírus da Encefalite Japonesa (Espécie)/patogenicidade , Encefalite Japonesa/patologia , Interações Hospedeiro-Patógeno/genética , Replicação Viral , Animais , Sistemas CRISPR-Cas/genética , Calreticulina/genética , Calreticulina/metabolismo , Vírus da Encefalite Japonesa (Espécie)/metabolismo , Encefalite Japonesa/virologia , Técnicas de Silenciamento de Genes , Técnicas de Inativação de Genes , Biblioteca Gênica , Células HEK293 , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , RNA Guia/genética , RNA Interferente Pequeno/metabolismo , Sus scrofa
12.
Int J Biol Sci ; 16(15): 3028-3036, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33061814

RESUMO

Coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2, with acute respiratory failure as the most significant symptom, has led to a global pandemic. Angiotensin-converting enzyme 2 (ACE2) is considered as the most important receptor of SARS-CoV-2 and wildly expressed in human tissues. Whereas, the extremely low expression of ACE2 in lung could hardly interpret the severe symptom of pneumonia in COVID-19 patients. Here we profiled two SARS-CoV-2 infection related genes, the transmembrane serine protease 2 (TMPRSS2) and the interferon-inducible transmembrane protein 3 (IFITM3), in human tissues and organs. Consistent with the expression and distribution of ACE2, TMPRSS2 was also highly expressed in digestive, urinary and reproductive systems, but low expressed in lung. Notably, the anti-virus protein IFITM3 also expressed much lower in lung than other tissues, which might be related to the severe lung symptoms of COVID-19. In addition, the low expression of IFITM3 in immune cells suggested that SARS-CoV-2 might attack lymphocytes and induce the cytokine release syndrome (CRS). Furthermore, cancer patients were considered as more susceptible to SARS-CoV-2 infection. Our data supposed that fourteen types of tumors might have different susceptibility to the virus according to ACE2, TMPRSS2 and IFITM3 expression patterns. Interestingly the prognosis of six types of cancers including breast carcinoma (BRCA), lung adenocarcinoma (LUAD), uterine corpus endometrial carcinoma (UCEC), renal clear cell carcinoma (KIRC), prostate adenocarcinoma (PRAD), and hepatocellular carcinoma (LIHC) were closely related to these gene expressions. Our study explored the expression and distribution profiles of two potential novel molecules that might participate in SARS-CoV-2 infection and involved in immunity, which may provide a functional basis for preventing infection of SARS-CoV-2.


Assuntos
Regulação Neoplásica da Expressão Gênica , Proteínas de Membrana/fisiologia , Neoplasias/metabolismo , Proteínas de Ligação a RNA/fisiologia , Receptores Virais/fisiologia , Serina Endopeptidases/fisiologia , Betacoronavirus , Infecções por Coronavirus/genética , Infecções por Coronavirus/metabolismo , Análise Mutacional de DNA , Regulação da Expressão Gênica , Voluntários Saudáveis , Humanos , Proteínas de Membrana/genética , Neoplasias/diagnóstico , Neoplasias/genética , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/genética , Pneumonia Viral/metabolismo , Prognóstico , Proteínas de Ligação a RNA/genética , Receptores Virais/genética , Serina Endopeptidases/genética , Distribuição Tecidual
13.
Nucleic Acids Res ; 48(19): 10832-10847, 2020 11 04.
Artigo em Inglês | MEDLINE | ID: mdl-33045730

RESUMO

Horizontally acquired genes are typically regulated by ancestral regulators. This regulation enables expression of horizontally acquired genes to be coordinated with that of preexisting genes. Here, we report a singular example of the opposite regulation: a horizontally acquired gene that controls an ancestral regulator, thereby promoting bacterial virulence. We establish that the horizontally acquired regulatory gene ssrB is necessary to activate the ancestral regulatory system PhoP/PhoQ of Salmonella enterica serovar Typhimurium (S. Typhimurium) in mildly acidic pH, which S. Typhimurium experiences inside macrophages. SsrB promotes phoP transcription by binding upstream of the phoP promoter. SsrB also increases ugtL transcription by binding to the ugtL promoter region, where it overcomes gene silencing by the heat-stable nucleoid structuring protein H-NS, enhancing virulence. The largely non-pathogenic species S. bongori failed to activate PhoP/PhoQ in mildly acidic pH because it lacks both the ssrB gene and the SsrB binding site in the target promoter. Low Mg2+ activated PhoP/PhoQ in both S. bongori and ssrB-lacking S. Typhimurium, indicating that the SsrB requirement for PhoP/PhoQ activation is signal-dependent. By controlling the ancestral genome, horizontally acquired genes are responsible for more crucial abilities, including virulence, than currently thought.


Assuntos
Proteínas de Bactérias/genética , Transferência Genética Horizontal , Proteínas de Membrana/genética , Salmonella typhimurium/genética , Fatores de Transcrição/genética , Animais , Proteínas de Bactérias/metabolismo , Linhagem Celular , Evolução Molecular , Feminino , Regulação Bacteriana da Expressão Gênica , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Regiões Promotoras Genéticas , Salmonella typhimurium/patogenicidade , Fatores de Transcrição/metabolismo , Ativação Transcricional , Virulência/genética
14.
Nat Commun ; 11(1): 5259, 2020 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-33067454

RESUMO

To increase understanding of the genomic landscape of acral melanoma, a rare form of melanoma occurring on palms, soles or nail beds, whole genome sequencing of 87 tumors with matching transcriptome sequencing for 63 tumors was performed. Here we report that mutational signature analysis reveals a subset of tumors, mostly subungual, with an ultraviolet radiation signature. Significantly mutated genes are BRAF, NRAS, NF1, NOTCH2, PTEN and TYRP1. Mutations and amplification of KIT are also common. Structural rearrangement and copy number signatures show that whole genome duplication, aneuploidy and complex rearrangements are common. Complex rearrangements occur recurrently and are associated with amplification of TERT, CDK4, MDM2, CCND1, PAK1 and GAB2, indicating potential therapeutic options.


Assuntos
Melanoma/genética , Neoplasias Cutâneas/genética , Feminino , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/metabolismo , Amplificação de Genes , Dosagem de Genes , Genômica , Humanos , Masculino , Melanoma/metabolismo , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mutação , Oxirredutases/genética , Oxirredutases/metabolismo , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Receptor Notch2/genética , Receptor Notch2/metabolismo , Neoplasias Cutâneas/metabolismo , Sequenciamento Completo do Genoma
15.
Phytomedicine ; 79: 153350, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33002827

RESUMO

BACKGROUND: Vascular endothelial activation is pivotal for the pathological development of various infectious and inflammatory diseases. Therapeutic interventions to prevent endothelial activation are of great clinical significance to achieve anti-inflammatory strategy. Previous studies indicate that the total flavonoids from the endemic herbal medicine Nervilia fordii (Hance) Schltr exerts potent anti-inflammatory effect and protective effect against endotoxin lipopolysaccharide (LPS)-induced acute lung injury, and shows clinical benefit in severe acute respiratory syndromes (SARS). However, the exact effective component of Nervilia fordii and its potential mechanism remain unknown. PURPOSE: The aim of this study was to investigate the effect and mechanism of rhamnocitrin (RH), a flavonoid extracted from Nervilia fordii, on LPS-induced endothelial activation. METHODS: The in vitro endothelial cell activation model was induced by LPS in human umbilical vein endothelial cells (HUVECs). Cell viability was measured to determine the cytotoxicity of RH. RT-PCR, Western blot, fluorescent probe and immunofluorescence were conducted to evaluate the effect and mechanism of RH against endothelial activation. RESULTS: RH was extracted and isolated from Nervilia fordii. RH at the concentration from 10-7 M-10-5 M inhibited the expressions of interlukin-6 (IL-6) and -8 (IL-8), monocyte chemotactic protein-1 (MCP-1), intercellular adhesion molecule-1 (ICAM-1), vascular cell-adhesion molecule-1 (VCAM-1), and plasminogen activator inhibitor-1 (PAI-1) in response to LPS challenge. Mechanistically, RH repressed calcium store-operated Ca2+ entry (SOCE) induced by LPS, which is due to downregulation of stromal interaction molecule-1 (STIM-1) following upregulating microRNA-185 (miR-185). Ultimately, RH abrogated LPS-induced activation of SOCE-mediated calcineurin/NFATc3 (nuclear factor of activated T cells, cytoplasmic 3) signaling pathway. CONCLUSION: The present study identifies RH as a potent inhibitor of endothelial activation. Since vascular endothelial activation is a pivotal cause of excessive cytokine production, leading to cytokine storm and severe pathology in infectious diseases such as SARS and the ongoing COVID-19 pneumonia disease, RH might suggest promising therapeutic potential in the management of cytokine storm in these diseases.


Assuntos
Endotélio Vascular/efeitos dos fármacos , Proteínas Sensoras de Cálcio Intracelular/metabolismo , Quempferóis/farmacologia , Proteínas de Membrana/metabolismo , Fatores de Transcrição NFATC/metabolismo , Proteínas de Neoplasias/metabolismo , Orchidaceae/química , Molécula 1 de Interação Estromal/metabolismo , Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/metabolismo , Infecções por Coronavirus/virologia , Endotélio Vascular/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Quempferóis/isolamento & purificação , Lipopolissacarídeos/farmacologia , Pandemias , Pneumonia Viral/metabolismo , Pneumonia Viral/virologia , Molécula 1 de Adesão de Célula Vascular/metabolismo
16.
Nat Commun ; 11(1): 5080, 2020 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-33033258

RESUMO

Natural transformation is the process by which bacteria take up genetic material from their environment and integrate it into their genome by homologous recombination. It represents one mode of horizontal gene transfer and contributes to the spread of traits like antibiotic resistance. In Vibrio cholerae, a type IVa pilus (T4aP) is thought to facilitate natural transformation by extending from the cell surface, binding to exogenous DNA, and retracting to thread this DNA through the outer membrane secretin, PilQ. Here, we use a functional tagged allele of VcPilQ purified from native V. cholerae cells to determine the cryoEM structure of the VcPilQ secretin in amphipol to ~2.7 Å. We use bioinformatics to examine the domain architecture and gene neighborhood of T4aP secretins in Proteobacteria in comparison with VcPilQ. This structure highlights differences in the architecture of the T4aP secretin from the type II and type III secretion system secretins. Based on our cryoEM structure, we design a series of mutants to reversibly regulate VcPilQ gate dynamics. These experiments support the idea of VcPilQ as a potential druggable target and provide insight into the channel that DNA likely traverses to promote the spread of antibiotic resistance via horizontal gene transfer by natural transformation.


Assuntos
Sistemas de Secreção Bacterianos/ultraestrutura , Microscopia Crioeletrônica , Fímbrias Bacterianas/ultraestrutura , Secretina/química , Vibrio cholerae/metabolismo , Vibrio cholerae/ultraestrutura , Cisteína/genética , Proteínas de Membrana/ultraestrutura , Modelos Moleculares , Mutação/genética , Filogenia , Domínios Proteicos , Transformação Bacteriana
17.
Nat Commun ; 11(1): 4909, 2020 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-32999291

RESUMO

Effectively activating macrophages against cancer is promising but challenging. In particular, cancer cells express CD47, a 'don't eat me' signal that interacts with signal regulatory protein alpha (SIRPα) on macrophages to prevent phagocytosis. Also, cancer cells secrete stimulating factors, which polarize tumor-associated macrophages from an antitumor M1 phenotype to a tumorigenic M2 phenotype. Here, we report that hybrid cell membrane nanovesicles (known as hNVs) displaying SIRPα variants with significantly increased affinity to CD47 and containing M2-to-M1 repolarization signals can disable both mechanisms. The hNVs block CD47-SIRPα signaling axis while promoting M2-to-M1 repolarization within tumor microenvironment, significantly preventing both local recurrence and distant metastasis in malignant melanoma models. Furthermore, by loading a stimulator of interferon genes (STING) agonist, hNVs lead to potent tumor inhibition in a poorly immunogenic triple negative breast cancer model. hNVs are safe, stable, drug loadable, and suitable for genetic editing. These properties, combined with the capabilities inherited from source cells, make hNVs an attractive immunotherapy.


Assuntos
Micropartículas Derivadas de Células/imunologia , Imunoterapia/métodos , Macrófagos/imunologia , Melanoma/terapia , Recidiva Local de Neoplasia/prevenção & controle , Neoplasias de Mama Triplo Negativas/terapia , Animais , Antígeno CD47/metabolismo , Linhagem Celular Tumoral/transplante , Modelos Animais de Doenças , Feminino , Células HEK293 , Humanos , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Melanoma/imunologia , Melanoma/secundário , Proteínas de Membrana/agonistas , Proteínas de Membrana/imunologia , Camundongos , Nanopartículas/administração & dosagem , Recidiva Local de Neoplasia/imunologia , Nucleotídeos Cíclicos/administração & dosagem , Receptores Imunológicos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , Neoplasias de Mama Triplo Negativas/imunologia , Evasão Tumoral/efeitos dos fármacos , Evasão Tumoral/imunologia , Microambiente Tumoral/imunologia
18.
Nat Commun ; 11(1): 4913, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-33004813

RESUMO

Reprograming of proline metabolism is critical for tumor growth. Here we show that PINCH-1 is highly expressed in lung adenocarcinoma and promotes proline synthesis through regulation of mitochondrial dynamics. Knockout (KO) of PINCH-1 increases dynamin-related protein 1 (DRP1) expression and mitochondrial fragmentation, which suppresses kindlin-2 mitochondrial translocation and interaction with pyrroline-5-carboxylate reductase 1 (PYCR1), resulting in inhibition of proline synthesis and cell proliferation. Depletion of DRP1 reverses PINCH-1 deficiency-induced defects on mitochondrial dynamics, proline synthesis and cell proliferation. Furthermore, overexpression of PYCR1 in PINCH-1 KO cells restores proline synthesis and cell proliferation, and suppresses DRP1 expression and mitochondrial fragmentation. Finally, ablation of PINCH-1 from lung adenocarcinoma in mouse increases DRP1 expression and inhibits PYCR1 expression, proline synthesis, fibrosis and tumor growth. Our results identify a signaling axis consisting of PINCH-1, DRP1 and PYCR1 that regulates mitochondrial dynamics and proline synthesis, and suggest an attractive strategy for alleviation of tumor growth.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adenocarcinoma de Pulmão/patologia , Proteínas com Domínio LIM/metabolismo , Neoplasias Pulmonares/patologia , Proteínas de Membrana/metabolismo , Células A549 , Proteínas Adaptadoras de Transdução de Sinal/genética , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/mortalidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Proliferação de Células/genética , Proteínas do Citoesqueleto/metabolismo , Modelos Animais de Doenças , Dinaminas/metabolismo , Feminino , Técnicas de Inativação de Genes , Humanos , Proteínas com Domínio LIM/genética , Pulmão/citologia , Pulmão/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidade , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Mitocôndrias/metabolismo , Dinâmica Mitocondrial , Proteínas Musculares/metabolismo , Proteínas de Neoplasias/metabolismo , Prolina/biossíntese , Proteínas Proto-Oncogênicas p21(ras)/genética , Pirrolina Carboxilato Redutases/metabolismo , Análise de Sobrevida
19.
Nat Commun ; 11(1): 5065, 2020 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-33033237

RESUMO

The type VI protein secretion system (T6SS) is a powerful needle-like machinery found in Gram-negative bacteria that can penetrate the cytosol of receiving cells in milliseconds by physical force. Anchored by its membrane-spanning complex (MC) and a baseplate (BP), the T6SS sheath-tube is assembled in a stepwise process primed by TssA and terminated by TagA. However, the molecular details of its assembly remain elusive. Here, we systematically examined the initiation and termination of contractile and non-contractile T6SS sheaths in MC-BP, tssA and tagA mutants by fluorescence microscopy. We observe long pole-to-pole sheath-tube structures in the non-contractile MC-BP defective mutants but not in the Hcp tube or VgrG spike mutants. Combining overexpression and genetic mutation data, we demonstrate complex effects of TssM, TssA and TagA interactions on T6SS sheath-tube dynamics. We also report promiscuous interactions of TagA with multiple T6SS components, similar to TssA. Our results demonstrate that priming of the T6SS sheath-tube assembly is not dependent on TssA, nor is the assembly termination dependent on the distal end TssA-TagA interaction, and highlight the tripartite control of TssA-TssM-TagA on sheath-tube initiation and termination.


Assuntos
Proteínas de Bactérias/metabolismo , Sistemas de Secreção Tipo VI/metabolismo , Vibrio cholerae/metabolismo , Proteínas de Bactérias/química , Membrana Celular/metabolismo , Escherichia coli/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Membrana/metabolismo , Viabilidade Microbiana , Modelos Biológicos , Mutação/genética , Ligação Proteica , Domínios Proteicos
20.
Nat Commun ; 11(1): 5499, 2020 10 30.
Artigo em Inglês | MEDLINE | ID: mdl-33127892

RESUMO

The epiblast, which provides the foundation of the future body, is actively reshaped during early embryogenesis, but the reshaping mechanisms are poorly understood. Here, using a 3D in vitro model of early epiblast development, we identify the canonical Wnt/ß-catenin pathway and its central downstream factor Esrrb as the key signalling cascade regulating the tissue-scale organization of the murine pluripotent lineage. Although in vivo the Wnt/ß-catenin/Esrrb circuit is dispensable for embryonic development before implantation, autocrine Wnt activity controls the morphogenesis and long-term maintenance of the epiblast when development is put on hold during diapause. During this phase, the progressive changes in the epiblast architecture and Wnt signalling response show that diapause is not a stasis but instead is a dynamic process with underlying mechanisms that can appear redundant during transient embryogenesis.


Assuntos
Diapausa/fisiologia , Células-Tronco Embrionárias/metabolismo , Receptores Estrogênicos/metabolismo , Via de Sinalização Wnt/fisiologia , Animais , Desenvolvimento Embrionário , Feminino , Camadas Germinativas/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Morfogênese , Proteínas Serina-Treonina Quinases/metabolismo , Receptores Estrogênicos/genética , beta Catenina/genética
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