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1.
Chem Biol Interact ; 337: 109366, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33549581

RESUMO

Tripartite motif-containing protein 26 (TRIM26) is a member of the TRIM protein family and has been demonstrated to play crucial roles in several types of cancers. However, the biological role of TRIM26 in bladder cancer and the mechanism have not been studied. In this study, we investigated the expression of TRIM26 in bladder cancer tissues and their adjacent non-tumor tissues by Western blot and qRT-PCR. In vitro investigations were performed to assess the roles of TRIM26 in bladder cancer using TRIM26-silencing and TRIM26-overexpressing bladder cancer cell lines. MTT and EdU assays were performed to evaluate cell proliferation. Cell migration and invasion were determined by transwell assays. Western blot analysis was performed to detect the expression levels of p-Akt, Akt, p-GSK3ß, GSK3ß, ß-catenin and c-Myc. Our results showed that TRIM26 expression was upregulated in human bladder cancer tissues and cell lines at both mRNA and protein levels. Knockdown of TRIM26 significantly inhibited the proliferation, migration and invasion of bladder cancer cells. In contrast, TRIM26 overexpression promoted bladder cancer cell proliferation, cell migration and invasion. Furthermore, knockdown of TRIM26 significantly decreased the levels of p-Akt, p-GSK3ß, ß-catenin and c-Myc in bladder cancer cells. Additionally, induction of Akt by SC79 treatment reversed the inhibitory effects of TRIM26 knockdown on the cellular behaviors of bladder cancer cells, while inhibition of ß-catenin reversed the effects of TRIM26 overexpression on the behaviors. Finally, knockdown of TRIM26 attenuated the growth of tumor xenografts in nude mice. In conclusion, these findings demonstrated that TRIM26 exerted an oncogenic role in bladder cancer through regulation of cell proliferation, migration and invasion via the Akt/GSK3ß/ß-catenin pathway.


Assuntos
Proliferação de Células , Glicogênio Sintase Quinase 3 beta/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Neoplasias da Bexiga Urinária/patologia , beta Catenina/metabolismo , Acetatos/farmacologia , Animais , Benzopiranos/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Humanos , Camundongos , Camundongos Nus , Proteínas Proto-Oncogênicas c-akt/agonistas , Interferência de RNA , RNA Interferente Pequeno/metabolismo , RNA Interferente Pequeno/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Transplante Heterólogo , Proteínas com Motivo Tripartido/antagonistas & inibidores , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/antagonistas & inibidores , Ubiquitina-Proteína Ligases/genética , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/metabolismo
2.
Life Sci ; 268: 118985, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33412211

RESUMO

The tripartite motif (TRIM) family is defined by the presence of a Really Interesting New Gene (RING) domain, one or two B-box motifs and a coiled-coil region. TRIM proteins play key roles in many biological processes, including innate immunity, tumorigenesis, cell differentiation and ontogenetic development. Alterations in TRIM gene and protein levels frequently emerge in a wide range of tumors and affect tumor progression. As canonical E3 ubiquitin ligases, TRIM proteins participate in ubiquitin-dependent proteolysis of prominent components of the p53, NF-κB and PI3K/AKT signaling pathways. The occurrence of ubiquitylation events induced by TRIM proteins sustains internal balance between tumor suppressive and tumor promoting genes. In this review, we summarized the diverse mechanism of TRIM proteins responsible for the most common malignancy, lung cancer. Furthermore, we also discussed recent progress in both the diagnosis and therapeutics of tumors contributed by TRIM proteins.


Assuntos
Antineoplásicos/farmacologia , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Biomarcadores Tumorais/metabolismo , Desenho de Fármacos , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Transdução de Sinais , Proteínas com Motivo Tripartido/antagonistas & inibidores , Proteínas com Motivo Tripartido/química , Proteínas com Motivo Tripartido/genética , Ubiquitinação
3.
Viruses ; 13(1)2021 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-33419081

RESUMO

Herpes simplex virus type 2 (HSV-2) is the primary cause of genital herpes which results in significant morbidity and mortality, especially in women, worldwide. HSV-2 is transmitted primarily through infection of epithelial cells at skin and mucosal surfaces. Our earlier work to examine interactions between HSV-2 and vaginal epithelial cells demonstrated that infection of the human vaginal epithelial cell line (VK2) with HSV-2 resulted in increased expression of TRIM26, a negative regulator of the Type I interferon pathway. Given that upregulation of TRIM26 could negatively affect anti-viral pathways, we decided to further study the role of TRIM26 in HSV-2 infection and replication. To do this, we designed and generated two cell lines derived from VK2s with TRIM26 overexpressed (OE) and knocked out (KO). Both, along with wildtype (WT) VK2, were infected with HSV-2 and viral titres were measured in supernatants 24 h later. Our results showed significantly enhanced virus production by TRIM26 OE cells, but very little replication in TRIM26 KO cells. We next examined interferon-ß production and expression of two distinct interferon stimulated genes (ISGs), MX1 and ISG15, in all three cell lines, prior to and following HSV-2 infection. The absence of TRIM26 (KO) significantly upregulated interferon-ß production at baseline and even further after HSV-2 infection. TRIM26 KO cells also showed significant increase in the expression of MX1 and ISG15 before and after HSV-2 infection. Immunofluorescent staining indicated that overexpression of TRIM26 substantially decreased the nuclear localization of IRF3, the primary mediator of ISG activation, before and after HSV-2 infection. Taken together, our data indicate that HSV-2 utilizes host factor TRIM26 to evade anti-viral response and thereby increase its replication in vaginal epithelial cells.


Assuntos
Células Epiteliais/virologia , Herpes Simples/genética , Herpesvirus Humano 2/fisiologia , Fator Regulador 3 de Interferon/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Células Cultivadas , Citocinas/metabolismo , Regulação para Baixo , Células Epiteliais/metabolismo , Técnicas de Inativação de Genes , Células HEK293 , Herpesvirus Humano 2/genética , Humanos , Fator Regulador 3 de Interferon/genética , Proteínas de Resistência a Myxovirus/metabolismo , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinas/metabolismo , Replicação Viral
4.
J Surg Res ; 257: 56-68, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32818785

RESUMO

BACKGROUND: Burn injuries (BIs) due to scalding are one of the most common accidents among children. BIs greater than 40% of total body surface area are considered extensive and result in local and systemic response. We sought to assess morphological and myogenic mechanisms through both short- and long-term intensive insulin therapies that affect the skeletal muscle after extensive skin BI in young rats. MATERIALS AND METHODS: Wistar rats aged 21 d were distributed into four groups: control (C), control with insulin (C + I), scald burn injury (SI), and SI with insulin (SI + I). The SI groups were submitted to a 45% total body surface area burn, and the C + I and SI + I groups received insulin (5 UI/Kg/d) for 4 or 14 d. Glucose tolerance and the homeostatic model assessment of insulin resistance index were determined. Gastrocnemius muscles were analyzed for histopathological, morphometric, and immunohistochemical myogenic parameters (Pax7, MyoD, and MyoG); in addition, the expression of genes related to muscle atrophy (MuRF1 and MAFbx) and its regulation (IGF-1) were also assessed. RESULTS: Short-term treatment with insulin favored muscle regeneration by primary myogenesis and decreased muscle atrophy in animals with BIs, whereas the long-term treatment modulated myogenesis by increasing the MyoD protein. Both treatments improved histopathological parameters and secondary myogenesis by increasing the MyoG protein. CONCLUSIONS: Treatment with insulin benefits myogenic parameters during regeneration and modulates MuRF1, an important mediator of muscle atrophy.


Assuntos
Queimaduras/complicações , Insulina/administração & dosagem , Desenvolvimento Muscular/efeitos dos fármacos , Atrofia Muscular/prevenção & controle , Animais , Glicemia/análise , Superfície Corporal , Queimaduras/patologia , Queimaduras/fisiopatologia , Expressão Gênica/efeitos dos fármacos , Fator de Crescimento Insulin-Like I/genética , Masculino , Proteínas Musculares/genética , Músculo Esquelético/química , Músculo Esquelético/patologia , Músculo Esquelético/fisiopatologia , Atrofia Muscular/etiologia , Atrofia Muscular/genética , Proteína MyoD/análise , Miogenina/análise , Fatores de Transcrição Box Pareados/análise , Ratos , Ratos Wistar , Proteínas Ligases SKP Culina F-Box/genética , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética
5.
Mol Cell ; 81(3): 599-613.e8, 2021 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-33373584

RESUMO

RNA helicases and E3 ubiquitin ligases mediate many critical functions in cells, but their actions have largely been studied in distinct biological contexts. Here, we uncover evolutionarily conserved rules of engagement between RNA helicases and tripartite motif (TRIM) E3 ligases that lead to their functional coordination in vertebrate innate immunity. Using cryoelectron microscopy and biochemistry, we show that RIG-I-like receptors (RLRs), viral RNA receptors with helicase domains, interact with their cognate TRIM/TRIM-like E3 ligases through similar epitopes in the helicase domains. Their interactions are avidity driven, restricting the actions of TRIM/TRIM-like proteins and consequent immune activation to RLR multimers. Mass spectrometry and phylogeny-guided biochemical analyses further reveal that similar rules of engagement may apply to diverse RNA helicases and TRIM/TRIM-like proteins. Our analyses suggest not only conserved substrates for TRIM proteins but also, unexpectedly, deep evolutionary connections between TRIM proteins and RNA helicases, linking ubiquitin and RNA biology throughout animal evolution.


Assuntos
Proteína DEAD-box 58/metabolismo , Imunidade Inata , Helicase IFIH1 Induzida por Interferon/metabolismo , Receptores Imunológicos/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Microscopia Crioeletrônica , Proteína DEAD-box 58/genética , Proteína DEAD-box 58/ultraestrutura , Epitopos , Evolução Molecular , Células HEK293 , Humanos , Helicase IFIH1 Induzida por Interferon/genética , Helicase IFIH1 Induzida por Interferon/ultraestrutura , Modelos Moleculares , Filogenia , Ligação Proteica , Conformação Proteica em alfa-Hélice , Domínios e Motivos de Interação entre Proteínas , Receptores Imunológicos/genética , Receptores Imunológicos/ultraestrutura , Proteínas com Motivo Tripartido/genética , Proteínas com Motivo Tripartido/ultraestrutura , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/ultraestrutura
6.
Viruses ; 13(1)2020 12 25.
Artigo em Inglês | MEDLINE | ID: mdl-33375604

RESUMO

Tripartite-motif-containing protein 5 isoform α (TRIM5α) is a cytoplasmic antiretroviral effector upregulated by type I interferons (IFN-I). We previously showed that two points mutations, R332G/R335G, in the retroviral capsid-binding region confer human TRIM5α the capacity to target and strongly restrict HIV-1 upon overexpression of the mutated protein. Here, we used clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9-mediated homology-directed repair (HDR) to introduce these two mutations in the endogenous human TRIM5 gene. We found 6 out of 47 isolated cell clones containing at least one HDR-edited allele. One clone (clone 6) had both alleles containing R332G, but only one of the two alleles containing R335G. Upon challenge with an HIV-1 vector, clone 6 was significantly less permissive compared to unmodified cells, whereas the cell clones with monoallelic modifications were only slightly less permissive. Following interferon (IFN)-ß treatment, inhibition of HIV-1 infection in clone 6 was significantly enhanced (~40-fold inhibition). TRIM5α knockdown confirmed that HIV-1 was inhibited by the edited TRIM5 gene products. Quantification of HIV-1 reverse transcription products showed that inhibition occurred through the expected mechanism. In conclusion, we demonstrate the feasibility of potently inhibiting a viral infection through the editing of innate effector genes. Our results also emphasize the importance of biallelic modification in order to reach significant levels of inhibition by TRIM5α.


Assuntos
Edição de Genes , Infecções por HIV/genética , Infecções por HIV/virologia , HIV-1/fisiologia , Linfócitos T/metabolismo , Linfócitos T/virologia , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Tropismo Viral/genética , Sistemas CRISPR-Cas , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Interações Hospedeiro-Patógeno/genética , Humanos , Células Jurkat , RNA Guia , Linfócitos T/imunologia
7.
Nat Commun ; 11(1): 5127, 2020 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-33046716

RESUMO

Despite the importance of AKT overactivation in tumor progression, results from clinical trials of various AKT inhibitors remain suboptimal, suggesting that AKT-driven tumor metastasis needs to be further understood. Herein, based on long non-coding RNA (lncRNA) profiling induced by active AKT, we identify that VAL (Vimentin associated lncRNA, LINC01546), which is directly induced by AKT/STAT3 signaling, functions as a potent pro-metastatic molecule and is essential for active AKT-induced tumor invasion, metastasis and anoikis resistance in lung adenocarcinoma (LAD). Impressively, chemosynthetic siRNAs against VAL shows great therapeutic potential in AKT overactivation-driven metastasis. Interestingly, similar to activated AKT in LAD cells, although unable to induce epithelial-mesenchymal transition (EMT), VAL exerts potent pro-invasive and pro-metastatic effects through directly binding to Vimentin and competitively abrogating Trim16-depedent Vimentin polyubiquitination and degradation. Taken together, our study provides an interesting demonstration of a lncRNA-mediated mechanism for active AKT-driven EMT-independent LAD metastasis and indicates the great potential of targeting VAL or Vimentin stability as a therapeutic approach.


Assuntos
Adenocarcinoma de Pulmão/metabolismo , Transição Epitelial-Mesenquimal , Neoplasias Pulmonares/metabolismo , RNA Longo não Codificante/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Vimentina/metabolismo , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/patologia , Adenocarcinoma de Pulmão/fisiopatologia , Animais , Linhagem Celular Tumoral , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/fisiopatologia , Camundongos , Metástase Neoplásica , Proteólise , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Longo não Codificante/genética , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação , Vimentina/genética
8.
PLoS Pathog ; 16(9): e1008844, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32886716

RESUMO

The genomes of RNA and small DNA viruses of vertebrates display significant suppression of CpG dinucleotide frequencies. Artificially increasing dinucleotide frequencies results in substantial attenuation of virus replication, suggesting that these compositional changes may facilitate recognition of non-self RNA sequences. Recently, the interferon inducible protein ZAP, was identified as the host factor responsible for sensing CpG in viral RNA, through direct binding and possibly downstream targeting for degradation. Using an arrayed interferon stimulated gene expression library screen, we identified ZAPS, and its associated factor TRIM25, as inhibitors of human cytomegalovirus (HCMV) replication. Exogenous expression of ZAPS and TRIM25 significantly reduced virus replication while knockdown resulted in increased virus replication. HCMV displays a strikingly heterogeneous pattern of CpG representation with specific suppression of CpG motifs within the IE1 major immediate early transcript which is absent in subsequently expressed genes. We demonstrated that suppression of CpG dinucleotides in the IE1 gene allows evasion of inhibitory effects of ZAP. We show that acute virus replication is mutually exclusive with high levels of cellular ZAP, potentially explaining the higher levels of CpG in viral genes expressed subsequent to IE1 due to the loss of pressure from ZAP in infected cells. Finally, we show that TRIM25 regulates alternative splicing between the ZAP short and long isoforms during HCMV infection and interferon induction, with knockdown of TRIM25 resulting in decreased ZAPS and corresponding increased ZAPL expression. These results demonstrate for the first time that ZAP is a potent host restriction factor against large DNA viruses and that HCMV evades ZAP detection through suppression of CpG dinucleotides within the major immediate early 1 transcript. Furthermore, TRIM25 is required for efficient upregulation of the interferon inducible short isoform of ZAP through regulation of alternative splicing.


Assuntos
Processamento Alternativo , Ilhas de CpG , Infecções por Citomegalovirus/metabolismo , Citomegalovirus/fisiologia , Regulação Viral da Expressão Gênica , Proteínas de Ligação a RNA/metabolismo , Proteínas Repressoras/metabolismo , Replicação Viral , Linhagem Celular , Infecções por Citomegalovirus/genética , Infecções por Citomegalovirus/patologia , Humanos , Proteínas Imediatamente Precoces , Proteínas de Ligação a RNA/genética , Proteínas Repressoras/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas com Motivo Tripartido/genética , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
9.
Proc Natl Acad Sci U S A ; 117(30): 17965-17976, 2020 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-32651277

RESUMO

Mobile genetic elements have significantly shaped our genomic landscape. LINE-1 retroelements are the only autonomously active elements left in the human genome. Since new insertions can have detrimental consequences, cells need to efficiently control LINE-1 retrotransposition. Here, we demonstrate that the intrinsic immune factor TRIM5α senses and restricts LINE-1 retroelements. Previously, rhesus TRIM5α has been shown to efficiently block HIV-1 replication, while human TRIM5α was found to be less active. Surprisingly, we found that both human and rhesus TRIM5α efficiently repress human LINE-1 retrotransposition. TRIM5α interacts with LINE-1 ribonucleoprotein complexes in the cytoplasm, which is essential for restriction. In line with its postulated role as pattern recognition receptor, we show that TRIM5α also induces innate immune signaling upon interaction with LINE-1 ribonucleoprotein complexes. The signaling events activate the transcription factors AP-1 and NF-κB, leading to the down-regulation of LINE-1 promoter activity. Together, our findings identify LINE-1 as important target of human TRIM5α, which restricts and senses LINE-1 via two distinct mechanisms. Our results corroborate TRIM5α as pattern recognition receptor and shed light on its previously undescribed activity against mobile genetic elements, such as LINE-1, to protect the integrity of our genome.


Assuntos
Elementos Nucleotídeos Longos e Dispersos , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Expressão Gênica , Genes Reporter , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata/genética , Macaca mulatta , Regiões Promotoras Genéticas , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Transporte Proteico , Transdução de Sinais , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética
10.
Nat Commun ; 11(1): 3624, 2020 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-32681036

RESUMO

TRIM family proteins play integral roles in the innate immune response to virus infection. MG53 (TRIM72) is essential for cell membrane repair and is believed to be a muscle-specific TRIM protein. Here we show human macrophages express MG53, and MG53 protein expression is reduced following virus infection. Knockdown of MG53 in macrophages leads to increases in type I interferon (IFN) upon infection. MG53 knockout mice infected with influenza virus show comparable influenza virus titres to wild type mice, but display increased morbidity accompanied by more accumulation of CD45+ cells and elevation of IFNß in the lung. We find that MG53 knockdown results in activation of NFκB signalling, which is linked to an increase in intracellular calcium oscillation mediated by ryanodine receptor (RyR). MG53 inhibits IFNß induction in an RyR-dependent manner. This study establishes MG53 as a new target for control of virus-induced morbidity and tissue injury.


Assuntos
Influenza Humana/imunologia , Interferon beta/metabolismo , Proteínas de Membrana/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Animais , Sinalização do Cálcio/imunologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Técnicas de Silenciamento de Genes , Técnicas de Inativação de Genes , Humanos , Imunidade Inata , Vírus da Influenza A Subtipo H1N1/imunologia , Influenza Humana/virologia , Interferon beta/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , NF-kappa B/metabolismo , RNA Interferente Pequeno , Transdução de Sinais/imunologia , Proteínas com Motivo Tripartido/genética
11.
PLoS Pathog ; 16(6): e1008611, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32511263

RESUMO

Human infection with avian influenza A (H5N1) and (H7N9) viruses causes severe respiratory diseases. PB1-F2 protein is a critical virulence factor that suppresses early type I interferon response, but the mechanism of its action in relation to high pathogenicity is not well understood. Here we show that PB1-F2 protein of H7N9 virus is a particularly potent suppressor of antiviral signaling through formation of protein aggregates on mitochondria and inhibition of TRIM31-MAVS interaction, leading to prevention of K63-polyubiquitination and aggregation of MAVS. Unaggregated MAVS accumulated on fragmented mitochondria is prone to degradation by both proteasomal and lysosomal pathways. These properties are proprietary to PB1-F2 of H7N9 virus but not shared by its counterpart in WSN virus. A recombinant virus deficient of PB1-F2 of H7N9 induces more interferon ß in infected cells. Our findings reveal a subtype-specific mechanism for destabilization of MAVS and suppression of interferon response by PB1-F2 of H7N9 virus.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Subtipo H7N9 do Vírus da Influenza A/metabolismo , Influenza Humana/metabolismo , Agregação Patológica de Proteínas/metabolismo , Transdução de Sinais , Proteínas Virais/metabolismo , Células A549 , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Cães , Células HEK293 , Humanos , Subtipo H7N9 do Vírus da Influenza A/genética , Influenza Humana/genética , Influenza Humana/patologia , Interferon beta/genética , Interferon beta/metabolismo , Células Madin Darby de Rim Canino , Mitocôndrias/genética , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Agregação Patológica de Proteínas/genética , Células THP-1 , Proteínas com Motivo Tripartido/genética , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Proteínas Virais/genética
12.
Mol Genet Genomics ; 295(5): 1281-1294, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32564135

RESUMO

The tripartite motif (TRIM) gene family encodes diverse distinct proteins that play important roles in many biological processes. However, the molecular evolution and phylogenetic relationships of TRIM genes in primates are still elusive. We performed a genomic approach to identify and characterize TRIM genes in human and other six primate genomes. In total, 537 putative functional TRIM genes were identified and TRIM members varied among primates. A neighbor joining (NJ) tree based on the protein sequences of 82 human TRIM genes indicates seven TRIM groups, which is consistent with the results based on the architectural motifs. Many TRIM gene duplication events were identified, indicating a recent expansion of TRIM family in primate lineages. Interestingly, the chimpanzee genome shows the greatest TRIM gene expansion among the primates; however, its congeneric species, bonobo, has the least number of TRIM genes and no duplication event. Moreover, we identified a ~ 200 kb deletion on chromosome 11 of bonobos that results in a loss of cluster3 TRIM genes. The loss of TRIM genes might have occurred within the last 2 mys. Analysis of positive selection recovered 9 previously reported and 21 newly identified positively selected TRIM genes. In particular, most positive selected sites are located in the B30.2 domains. Our results have provided new insight into the evolution of primate TRIM genes and will broaden our understanding on the functions of the TRIM family.


Assuntos
Genômica/métodos , Primatas/genética , Proteínas com Motivo Tripartido/genética , Animais , Evolução Molecular , Duplicação Gênica , Gorilla gorilla/genética , Humanos , Macaca/genética , Família Multigênica , Pan paniscus/genética , Pan troglodytes/genética , Filogenia , Seleção Genética , Deleção de Sequência
13.
Int J Infect Dis ; 97: 47-53, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32531432

RESUMO

PURPOSE: To explore the molecular genetic mechanisms underlying different responsiveness to Enterovirus 71 (EV71) vaccine. METHODS: We recruited 10,245 healthy children into a phase 3 clinical trial to evaluate the efficacy of EV71 vaccine in 2012. Fifty subjects from the trial were divided into the potent immune response group (20 subjects) and the ineffective immune response group (30 subjects). Whole-exome sequencing was performed for these 50 samples and we conducted bioinformatics analyses based on online public database. RESULTS: A total of 222,180 germline variants were detected across 50 subjects. Single nucleotide variant (SNV)-based screening of the subjects with potent or ineffective immune response allowed the identification of a potentially detrimental heterozygous missense variant (c.3784C>T) in EEA1. We also retained TRIM59 and ABCA7 genes that contain different loss of function (LoF) variants shared in two cases and involved in the immune response process. Then, we conducted high-resolution typing of 9 classical HLA genes, HLA-DRB1*03:01, HLA-DQA1*05:01 and HLA-DQB1*02:01 alleles were frequently (recurrence ≥5) observed only in ineffective immune responders. CONCLUSIONS: Our study is a meaningful attempt on the comparison of genomic profiles between potent and ineffective immune responders induced by EV71 vaccine, and several candidate potentially detrimental genes were identified.


Assuntos
Enterovirus Humano A/imunologia , Vacinas Virais/imunologia , Transportadores de Cassetes de Ligação de ATP/genética , Alelos , Pré-Escolar , China , Feminino , Variação Genética , Cadeias alfa de HLA-DQ/genética , Cadeias beta de HLA-DQ/genética , Cadeias HLA-DRB1/genética , Humanos , Lactente , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Proteínas com Motivo Tripartido/genética , Sequenciamento Completo do Exoma
14.
Proc Natl Acad Sci U S A ; 117(25): 14395-14404, 2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32513696

RESUMO

Retinoic acid-inducible gene I (RIG-I) is up-regulated during granulocytic differentiation of acute promyelocytic leukemia (APL) cells induced by all-trans retinoic acid (ATRA). It has been reported that RIG-I recognizes virus-specific 5'-ppp-double-stranded RNA (dsRNA) and activates the type I interferons signaling pathways in innate immunity. However, the functions of RIG-I in hematopoiesis remain unclear, especially regarding its possible interaction with endogenous RNAs and the associated pathways that could contribute to the cellular differentiation and maturation. Herein, we identified a number of RIG-I-binding endogenous RNAs in APL cells following ATRA treatment, including the tripartite motif-containing protein 25 (TRIM25) messenger RNA (mRNA). TRIM25 encodes the protein known as an E3 ligase for ubiquitin/interferon (IFN)-induced 15-kDa protein (ISG15) that is involved in RIG-I-mediated antiviral signaling. We show that RIG-I could bind TRIM25 mRNA via its helicase domain and C-terminal regulatory domain, enhancing the stability of TRIM25 transcripts. RIG-I could increase the transcriptional expression of TRIM25 by caspase recruitment domain (CARD) domain through an IFN-stimulated response element. In addition, RIG-I activated other key genes in the ISGylation pathway by activating signal transducer and activator of transcription 1 (STAT1), including the modifier ISG15 and several enzymes responsible for the conjugation of ISG15 to protein substrates. RIG-I cooperated with STAT1/2 and interferon regulatory factor 1 (IRF1) to promote the activation of the ISGylation pathway. The integrity of ISGylation in ATRA or RIG-I-induced cell differentiation was essential given that knockdown of TRIM25 or ISG15 resulted in significant inhibition of this process. Our results provide insight into the role of the RIG-I-TRIM25-ISGylation axis in myeloid differentiation.


Assuntos
Diferenciação Celular , Citocinas/metabolismo , Proteína DEAD-box 58/metabolismo , Granulócitos/fisiologia , Fatores de Transcrição/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinas/metabolismo , Linhagem Celular Tumoral , Citocinas/genética , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Estabilidade de RNA , RNA Mensageiro/metabolismo , Fatores de Transcrição/genética , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinas/genética , Regulação para Cima
15.
J Vet Sci ; 21(3): e49, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32476322

RESUMO

BACKGROUND: Coinfection with avian leukosis virus subgroup J (ALV-J) and reticuloendotheliosis virus (REV) is common in chickens, and the molecular mechanism of the synergistic pathogenic effects of the coinfection is not clear. Exosomes have been identified as new players in the pathogenesis of retroviruses. The different functions of exosomes depend on their cargo components. OBJECTIVES: The aim of this study was to investigate the function of co-regulation differentially expressed proteins in exosomes on coinfection of ALV-J and REV. METHODS: Here, viral replication in CEF cells infected with ALV-J, REV or both was detected by immunofluorescence microscopy. Then, we analyzed the exosomes isolated from supernatants of chicken embryo fibroblast (CEF) cells single infected and coinfected with ALV-J and REV by mass spectrometry. KEGG pathway enrichment analyzed the co-regulation differentially expressed proteins in exosomes. Next, we silenced and overexpressed tripartite motif containing 62 (TRIM62) to evaluate the effects of TRIM62 on viral replication and the expression levels of NCK-association proteins 1 (NCKAP1) and actin-related 2/3 complex subunit 5 (ARPC5) determined by quantitative reverse transcription polymerase chain reaction. RESULTS: The results showed that coinfection of ALV-J and REV promoted the replication of each other. Thirty proteins, including TRIM62, NCK-association proteins 1 (NCKAP1, also known as Nap125), and Arp2/3-5, ARPC5, were identified. NCKAP1 and ARPC5 were involved in the actin cytoskeleton pathway. TRIM62 negatively regulated viral replication and that the inhibition of REV was more significant than that on ALV-J in CEF cells coinfected with TRIM62. In addition, TRIM62 decreased the expression of NCKAP1 and increased the expression of ARPC5 in coinfected CEF cells. CONCLUSIONS: Collectively, our results indicated that coinfection with ALV-J and REV competitively promoted each other's replication, the actin cytoskeleton played an important role in the coinfection mechanism, and TRIM62 regulated the actin cytoskeleton.


Assuntos
Proteínas Aviárias/genética , Coinfecção/veterinária , Regulação da Expressão Gênica , Doenças das Aves Domésticas/fisiopatologia , Infecções por Retroviridae/veterinária , Proteínas com Motivo Tripartido/genética , Citoesqueleto de Actina/genética , Citoesqueleto de Actina/metabolismo , Animais , Leucose Aviária/fisiopatologia , Leucose Aviária/virologia , Vírus da Leucose Aviária/fisiologia , Proteínas Aviárias/metabolismo , Coinfecção/fisiopatologia , Coinfecção/virologia , Doenças das Aves Domésticas/virologia , Vírus da Reticuloendoteliose/fisiologia , Infecções por Retroviridae/fisiopatologia , Infecções por Retroviridae/virologia , Proteínas com Motivo Tripartido/metabolismo
16.
Adv Exp Med Biol ; 1233: 311-325, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32274764

RESUMO

The TRIM family comprises proteins characterized by the presence of the tripartite motif composed of a RING domain, one or two B-box domains and a coiled-coil region. The TRIM shared domain structure underscores a common biochemical function as E3 ligase within the ubiquitination cascade. The TRIM proteins represent one of the largest E3 ligase families counting in human more than 70 members. These proteins are implicated in a plethora of cellular processes such as apoptosis, cell cycle regulation, muscular physiology, and innate immune response. Consistently, their alteration results in several pathological conditions emphasizing their medical relevance. Here, the genetic and pathogenetic mechanisms of rare disorders directly caused by mutations in TRIM genes will be reviewed. These diseases fall into different pathological areas, from malformation birth defects due to developmental abnormalities, to neurological disorders and progressive teenage neuromuscular disorders. In many instances, TRIM E3 ligases act on several substrates thus exerting pleiotropic activities: the need of unraveling disease-specific TRIM pathways for a precise targeting therapy avoiding dramatic side effects will be discussed.


Assuntos
Doenças Genéticas Inatas/enzimologia , Doenças Genéticas Inatas/genética , Doenças Raras/enzimologia , Doenças Raras/genética , Proteínas com Motivo Tripartido/química , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/química , Ubiquitina-Proteína Ligases/metabolismo , Humanos , Domínios Proteicos , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
17.
Am J Physiol Cell Physiol ; 319(2): C392-C401, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32348176

RESUMO

This study was conducted to define the underlying molecular mechanism of tripartite motif (TRIM) 59-induced invasion of ectopic endometrial stromal cells in endometriosis. Primary endometriosis ectopic endometrial stromal cells and normal endometrial cells were isolated and purified. Western blot was used to detect the expression of TRIM59, protein phosphatase Mg2+/Mn2+-dependent 1A (PPM1A), smad2/3, and phosphorylated (p)-smad2/3. Lentiviral vector-mediated TRIM59 interference and overexpression were established. Cell Counting Kit-8 assay was used to detect cell proliferation, and the Transwell migration assay was used to detect cell invasion. Matrix metalloproteinase (MMP-2), MMP9, smad2/3, and p-smad2/3 expressions were also detected using Western blot analysis; degradation of PPM1A was verified to be through ubiquitination. We found that TRIM59 expression levels in the endometriosis group was significantly higher compared with the normal group (P < 0.05), whereas the expression levels of PPM1A in the endometriosis group were significantly lower (P < 0.05). Endometriosis did not alter smad2/3 (P > 0.05) expression. However, after activating smad2/3 by phosphorylation, the expression of p-smad2/3 in the endometriosis group was significantly higher compared with the normal group (P < 0.05). The content of PPM1A in the TRIM59 overexpression group was significantly lower than that in the control group (P < 0.001), whereas the content of PPM1A in the siTRIM59 group was significantly higher than that in the control group (P < 0.001). In addition, there were no significant differences in the mRNA levels of PPM1A among the five groups, indicating that TRIM59 affects the expression of PPM1A at the posttranslational level (P < 0.05). Overexpression of TRIM59 significantly promoted the ubiquitination of PPM1A. We conclude that TRIM59 inhibits PPM1A through ubiquitination and activates the transforming growth factor-ß/Smad pathway to promote the invasion of ectopic endometrial stromal cells in endometriosis.


Assuntos
Endometriose/genética , Endométrio/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteína Fosfatase 2C/genética , Fator de Crescimento Transformador beta/genética , Proteínas com Motivo Tripartido/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Expressão Ectópica do Gene/genética , Endometriose/patologia , Endométrio/patologia , Feminino , Humanos , Cultura Primária de Células , Transdução de Sinais/genética , Proteína Smad2/genética , Células Estromais/metabolismo , Células Estromais/patologia , Ubiquitinação/genética
18.
PLoS Pathog ; 16(3): e1008387, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32126128

RESUMO

Mediator of IRF3 activation (MITA, also named as STING/ERIS/MPYS/TMEM173), is essential to DNA virus- or cytosolic DNA-triggered innate immune responses. In this study, we demonstrated the negative regulatory role of RING-finger protein (RNF) 90 in innate immune responses targeting MITA. RNF90 promoted K48-linked ubiquitination of MITA and its proteasome-dependent degradation. Overexpression of RNF90 inhibited HSV-1- or cytosolic DNA-induced immune responses whereas RNF90 knockdown had the opposite effects. Moreover, RNF90-deficient bone marrow-derived dendritic cells (BMDCs), bone marrow-derived macrophages (BMMs) and mouse embryonic fibroblasts (MEFs) exhibited increased DNA virus- or cytosolic DNA-triggered signaling and RNF90 deficiency protected mice from DNA virus infection. Taken together, our findings suggested a novel function of RNF90 in innate immunity.


Assuntos
Herpesvirus Humano 1/imunologia , Imunidade Inata , Proteínas de Membrana/imunologia , Proteólise , Proteínas com Motivo Tripartido/imunologia , Ubiquitina-Proteína Ligases/imunologia , Ubiquitinação/imunologia , Animais , Células da Medula Óssea/imunologia , Células da Medula Óssea/virologia , Células Dendríticas/imunologia , Células Dendríticas/virologia , Fibroblastos/imunologia , Fibroblastos/virologia , Herpesvirus Humano 1/genética , Macrófagos/imunologia , Macrófagos/virologia , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação/genética
19.
Scand J Immunol ; 91(6): e12876, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32145086

RESUMO

Rheumatoid arthritis (RA) is a worldwide autoimmune disease. The study of its aetiology and mechanism has always been a focus topic in medicine. This research was designed to investigate the effect of E3 ubiquitin ligase tripartite motif protein 32 (TRIM32) in rheumatoid arthritis (RA). We found in fibroblast-like synoviocytes (FLS) of RA patients, the expression of TRIM32 was significantly increased compared with its expression in osteoarthritis (OA) patients FLS. A widely used pro-inflammatory stimuli tumour necrosis factor-alpha (TNF-α) was found to promote TRIM32 expression in a time-dependent manner. Furthermore, we observed that overexpression of TRIM32 aggravated the production of pro-inflammatory cytokines in FLS, silencing of TRIM32 showed the consistent results. In addition, TRIM32 was found to activate nuclear factor κB (NF-κB) signalling pathway, and TRIM32 could interact with TNF receptor-associated factor 2 (TRAF2) to promote the K63-linked polyubiquitination of TRAF2 in RA-FLS. In conclusion, we suggested that TRIM32 as a positive regulator of inflammatory responses in RA-FLS.


Assuntos
Artrite Reumatoide/metabolismo , Fibroblastos/imunologia , Inflamação/metabolismo , Sinoviócitos/imunologia , Fatores de Transcrição/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Artrite Reumatoide/imunologia , Proliferação de Células , Células Cultivadas , Citocinas/metabolismo , Humanos , Inflamação/imunologia , Mediadores da Inflamação/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais , Fator 2 Associado a Receptor de TNF/metabolismo , Fatores de Transcrição/genética , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação , Regulação para Cima
20.
Int J Mol Sci ; 21(3)2020 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-32050585

RESUMO

Myostatin is a myokine that regulates muscle function and mass, producing muscle atrophy. Myostatin induces the degradation of myofibrillar proteins, such as myosin heavy chain or troponin. The main pathway that mediates protein degradation during muscle atrophy is the ubiquitin proteasome system, by increasing the expression of atrogin-1 and MuRF-1. In addition, myostatin activates the NF-κB signaling pathway. Renin-angiotensin system (RAS) also regulates muscle mass. Angiotensin (1-7) (Ang-(1-7)) has anti-atrophic properties in skeletal muscle. In this paper, we evaluated the effect of Ang-(1-7) on muscle atrophy and signaling induced by myostatin. The results show that Ang-(1-7) prevented the decrease of the myotube diameter and myofibrillar protein levels induced by myostatin. Ang-(1-7) also abolished the increase of myostatin-induced reactive oxygen species production, atrogin-1, MuRF-1, and TNF-α gene expressions and NF-κB signaling activation. Ang-(1-7) inhibited the activity mediated by myostatin through Mas receptor, as is demonstrated by the loss of all Ang-(1-7)-induced effects when the Mas receptor antagonist A779 was used. Our results show that the effects of Ang-(1-7) on the myostatin-dependent muscle atrophy and signaling are blocked by MK-2206, an inhibitor of Akt/PKB. Together, these data indicate that Ang-(1-7) inhibited muscle atrophy and signaling induced by myostatin through a mechanism dependent on Mas receptor and Akt/PKB.


Assuntos
Angiotensina I/farmacologia , Fibras Musculares Esqueléticas/efeitos dos fármacos , Miostatina/farmacologia , NF-kappa B/metabolismo , Fragmentos de Peptídeos/farmacologia , Transdução de Sinais , Animais , Linhagem Celular , Camundongos , Fibras Musculares Esqueléticas/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Ligases SKP Culina F-Box/genética , Proteínas Ligases SKP Culina F-Box/metabolismo , Proteínas com Motivo Tripartido/genética , Proteínas com Motivo Tripartido/metabolismo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
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