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1.
Nat Commun ; 11(1): 4586, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32934222

RESUMO

Frequent mutation of the tumour suppressor RNF43 is observed in many cancers, particularly colon malignancies. RNF43, an E3 ubiquitin ligase, negatively regulates Wnt signalling by inducing degradation of the Wnt receptor Frizzled. In this study, we discover that RNF43 activity requires phosphorylation at a triplet of conserved serines. This phospho-regulation of RNF43 is required for zebrafish development and growth of mouse intestinal organoids. Cancer-associated mutations that abrogate RNF43 phosphorylation cooperate with active Ras to promote tumorigenesis by abolishing the inhibitory function of RNF43 in Wnt signalling while maintaining its inhibitory function in p53 signalling. Our data suggest that RNF43 mutations cooperate with KRAS mutations to promote multi-step tumorigenesis via the Wnt-Ras-p53 axis in human colon cancers. Lastly, phosphomimetic substitutions of the serine trio restored the tumour suppressive activity of extracellular oncogenic mutants. Therefore, harnessing phospho-regulation of RNF43 might be a potential therapeutic strategy for tumours with RNF43 mutations.


Assuntos
Carcinogênese/metabolismo , Receptores Wnt/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Carcinogênese/genética , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Proteína Oncogênica p21(ras)/genética , Proteína Oncogênica p21(ras)/metabolismo , Fosforilação , Proteólise , Receptores Wnt/genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína Ligases/genética , Via de Sinalização Wnt
2.
PLoS Comput Biol ; 16(9): e1007740, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32881861

RESUMO

The circadian clock is a complex system that plays many important roles in most organisms. Previously, many mathematical models have been used to sharpen our understanding of the Arabidopsis clock, which brought to light the roles of each transcriptional and post-translational regulations. However, the presence of both regulations, instead of either transcription or post-translation, raised curiosity of whether the combination of these two regulations is important for the clock's system. In this study, we built a series of simplified oscillators with different regulations to study the importance of post-translational regulation (specifically, 26S proteasome degradation) in the clock system. We found that a simple transcriptional-based oscillator can already generate sustained oscillation, but the oscillation can be easily destroyed in the presence of transcriptional leakage. Coupling post-translational control with transcriptional-based oscillator in a feed-forward loop will greatly improve the robustness of the oscillator in the presence of basal leakage. Using these general models, we were able to replicate the increased variability observed in the E3 ligase mutant for both plant and mammalian clocks. With this insight, we also predict a plausible regulator of several E3 ligase genes in the plant's clock. Thus, our results provide insights into and the plausible importance in coupling transcription and post-translation controls in the clock system.


Assuntos
Relógios Circadianos/genética , Modelos Biológicos , Processamento de Proteína Pós-Traducional/genética , Transcrição Genética/genética , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Biologia Computacional , Retroalimentação Fisiológica , Regulação da Expressão Gênica de Plantas/genética , Complexo de Endopeptidases do Proteassoma/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
3.
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
4.
Medicine (Baltimore) ; 99(35): e21989, 2020 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-32871949

RESUMO

BACKGROUND: Bipolar disorder (BD), a common kind of mood disorder with frequent recurrence, high rates of additional comorbid conditions and poor compliance, has an unclear pathogenesis. The Gene Expression Omnibus (GEO) database is a gene expression database created and maintained by the National Center for Biotechnology Information. Researchers can download expression data online for bioinformatics analysis, especially for cancer research. However, there is little research on the use of such bioinformatics analysis methodologies for mental illness by downloading differential expression data from the GEO database. METHODS: Publicly available data were downloaded from the GEO database (GSE12649, GSE5388 and GSE5389), and differentially expressed genes (DEGs) were extracted by using the online tool GEO2R. A Venn diagram was used to screen out common DEGs between postmortem brain tissues and normal tissues. Functional annotation and pathway enrichment analysis of DEGs were performed by using Gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses, respectively. Furthermore, a protein-protein interaction network was constructed to identify hub genes. RESULTS: A total of 289 DEGs were found, among which 5 of 10 hub genes [HSP90AA1, HSP90AB 1, UBE2N, UBE3A, and CUL1] were identified as susceptibility genes whose expression was downregulated. Gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses showed that variations in these 5 hub genes were obviously enriched in protein folding, protein polyubiquitination, apoptotic process, protein binding, the ubiquitin-mediated proteolysis pathway, and protein processing in the endoplasmic reticulum pathway. These findings strongly suggested that HSP90AA1, UBE3A, and CUL 1, which had large areas under the curve in receiver operator curves (P < .05), were potential diagnostic markers for BD. CONCLUSION: Although there are 3 hub genes [HSP90AA1, UBE3A, and CUL 1] that are tightly correlated with the occurrence of BD, mainly based on routine bioinformatics methods for cancer-related disease, the feasibility of applying this single GEO bioinformatics approach for mental illness is questionable, given the significant differences between mental illness and cancer-related diseases.


Assuntos
Transtorno Bipolar/genética , Proteínas Culina/genética , Mineração de Dados/métodos , Proteínas de Choque Térmico HSP90/genética , Ubiquitina-Proteína Ligases/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Feminino , Predisposição Genética para Doença , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Neoplasias/genética , Mapas de Interação de Proteínas , Adulto Jovem
5.
Sci Adv ; 6(33): eabb7238, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32851183

RESUMO

Cigarette smoking, the leading cause of chronic obstructive pulmonary disease (COPD), has been implicated as a risk factor for severe disease in patients infected with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here we show that mice with lung epithelial cell-specific loss of function of Miz1, which we identified as a negative regulator of nuclear factor κB (NF-κB) signaling, spontaneously develop progressive age-related changes resembling COPD. Furthermore, loss of Miz1 up-regulates the expression of Ace2, the receptor for SARS-CoV-2. Concomitant partial loss of NF-κB/RelA prevented the development of COPD-like phenotype in Miz1-deficient mice. Miz1 protein levels are reduced in the lungs from patients with COPD, and in the lungs of mice exposed to chronic cigarette smoke. Our data suggest that Miz1 down-regulation-induced sustained activation of NF-κB-dependent inflammation in the lung epithelium is sufficient to induce progressive lung and airway destruction that recapitulates features of COPD, with implications for COVID-19.


Assuntos
Células Epiteliais/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Pulmão/metabolismo , Peptidil Dipeptidase A/metabolismo , Fenótipo , Proteínas Inibidoras de STAT Ativados/genética , Doença Pulmonar Obstrutiva Crônica/genética , Ubiquitina-Proteína Ligases/genética , Regulação para Cima/genética , Animais , Betacoronavirus , Infecções por Coronavirus/metabolismo , Infecções por Coronavirus/virologia , Técnicas de Inativação de Genes , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pandemias , Pneumonia Viral/metabolismo , Pneumonia Viral/virologia , Proteínas Inibidoras de STAT Ativados/metabolismo , Doença Pulmonar Obstrutiva Crônica/etiologia , Doença Pulmonar Obstrutiva Crônica/metabolismo , Transdução de Sinais/genética , Fumar/efeitos adversos , Fator de Transcrição RelA/metabolismo , Ubiquitina-Proteína Ligases/metabolismo
6.
Nat Commun ; 11(1): 3807, 2020 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-32733036

RESUMO

The human genome contains an estimated 600 ubiquitin E3 ligases, many of which are single-subunit E3s (ssE3s) that can bind to both substrate and ubiquitin-loaded E2 (E2~Ub). Within ssE3s structural disorder tends to be located in substrate binding and domain linking regions. RNF4 is a ssE3 ligase with a C-terminal RING domain and disordered N-terminal region containing SUMO Interactions Motifs (SIMs) required to bind SUMO modified substrates. Here we show that, although the N-terminal region of RNF4 bears no secondary structure, it maintains a compact global architecture primed for SUMO interaction. Segregated charged regions within the RNF4 N-terminus promote compaction, juxtaposing RING domain and SIMs to facilitate substrate ubiquitination. Mutations that induce a more extended shape reduce ubiquitination activity. Our result offer insight into a key step in substrate ubiquitination by a member of the largest ubiquitin ligase subtype and reveal how a defined architecture within a disordered region contributes to E3 ligase function.


Assuntos
Proteínas Intrinsicamente Desordenadas/metabolismo , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Humanos , Proteínas Intrinsicamente Desordenadas/genética , Proteínas Nucleares/genética , Ligação Proteica , Domínios Proteicos , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismo , Fatores de Transcrição/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
7.
Nat Commun ; 11(1): 4205, 2020 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-32826891

RESUMO

Triple negative breast cancer (TNBC) is a deadly form of breast cancer due to the development of resistance to chemotherapy affecting over 30% of patients. New therapeutics and companion biomarkers are urgently needed. Recognizing the elevated expression of glucose transporter 1 (GLUT1, encoded by SLC2A1) and associated metabolic dependencies in TNBC, we investigated the vulnerability of TNBC cell lines and patient-derived samples to GLUT1 inhibition. We report that genetic or pharmacological inhibition of GLUT1 with BAY-876 impairs the growth of a subset of TNBC cells displaying high glycolytic and lower oxidative phosphorylation (OXPHOS) rates. Pathway enrichment analysis of gene expression data suggests that the functionality of the E2F pathway may reflect to some extent OXPHOS activity. Furthermore, the protein levels of retinoblastoma tumor suppressor (RB1) strongly correlate with the degree of sensitivity to GLUT1 inhibition in TNBC, where RB1-negative cells are insensitive to GLUT1 inhibition. Collectively, our results highlight a strong and targetable RB1-GLUT1 metabolic axis in TNBC and warrant clinical evaluation of GLUT1 inhibition in TNBC patients stratified according to RB1 protein expression levels.


Assuntos
Transportador de Glucose Tipo 1/antagonistas & inibidores , Transportador de Glucose Tipo 1/metabolismo , Proteínas de Ligação a Retinoblastoma/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Apoptose/efeitos dos fármacos , Biomarcadores Tumorais , Neoplasias da Mama/metabolismo , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Transportador de Glucose Tipo 1/genética , Humanos , Camundongos , Fosforilação Oxidativa , Proteômica , Pirazóis/farmacologia , Piridinas/farmacologia , Quinolinas , RNA Mensageiro/metabolismo , Neoplasias de Mama Triplo Negativas/genética , Ubiquitina-Proteína Ligases/genética
8.
Mol Cell ; 79(6): 950-962.e6, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32726578

RESUMO

Ribosome-associated quality control (RQC) pathways protect cells from toxicity caused by incomplete protein products resulting from translation of damaged or problematic mRNAs. Extensive work in yeast has identified highly conserved mechanisms that lead to degradation of faulty mRNA and partially synthesized polypeptides. Here we used CRISPR-Cas9-based screening to search for additional RQC strategies in mammals. We found that failed translation leads to specific inhibition of translation initiation on that message. This negative feedback loop is mediated by two translation inhibitors, GIGYF2 and 4EHP. Model substrates and growth-based assays established that inhibition of additional rounds of translation acts in concert with known RQC pathways to prevent buildup of toxic proteins. Inability to block translation of faulty mRNAs and subsequent accumulation of partially synthesized polypeptides could explain the neurodevelopmental and neuropsychiatric disorders observed in mice and humans with compromised GIGYF2 function.


Assuntos
Proteínas de Transporte/genética , Fator de Iniciação 4E em Eucariotos/genética , Iniciação Traducional da Cadeia Peptídica , Ribossomos/genética , Animais , Sistemas CRISPR-Cas/genética , Humanos , Camundongos , Biossíntese de Proteínas/genética , Processamento de Proteína Pós-Traducional/genética , Controle de Qualidade , RNA Mensageiro/genética , Ubiquitina-Proteína Ligases/genética
9.
Prostate ; 80(12): 1012-1023, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32649013

RESUMO

BACKGROUND: Small cell neuroendocrine (NE) carcinomas of the prostate classically lose androgen receptor (AR) expression, may harbor loss of the RB1, TP53, and PTEN tumor suppressor genes, and are associated with a poor prognosis. However usual-type adenocarcinomas may also contain areas of NE differentiation, and in this context the molecular features and biological significance are less certain. METHODS: We examined the molecular phenotype and oncologic outcomes of primary prostate adenocarcinomas with ≥5% NE differentiation (≥5% chromogranin A-positive NE cells in any given tumor spot on tissue microarray) using three independent study sets: a set of tumors with paneth cell-like NE differentiation (n = 26), a retrospective case-cohort of intermediate- and high-risk patients enriched for adverse outcomes (n = 267), and primary tumors from a retrospective series of men with eventual castration-resistant metastatic prostate cancer (CRPC) treated with abiraterone or enzalutamide (n = 55). RESULTS: Benign NE cells expressed significantly lower quantified AR levels compared with paired benign luminal cells (P < .001). Similarly, paneth-like NE carcinoma cells or carcinoma cells expressing chromogranin A expressed significantly lower quantified AR levels than paired non-NE carcinoma cells (P < .001). Quantified ERG protein expression, was also lower in chromogranin A-labeled adenocarcinoma cells compared with unlabeled cells (P < .001) and tumors with NE differentiation showed lower gene expression scores for AR activity compared with those without. Despite evidence of lower AR signaling, adenocarcinomas with NE differentiation did not differ by prevalence of TP53 missense mutations, or PTEN or RB1 loss, compared with those without NE differentiation. Finally, NE differentiation was not associated with time to metastasis in intermediate- and high-risk patients (P = .6 on multivariate analysis), nor with progression-free survival in patients with CRPC treated with abiraterone or enzalutamide (P = .9). CONCLUSION: NE differentiation in usual-type primary prostate adenocarcinoma is a molecularly and clinically distinct form of lineage plasticity from that occurring in small cell NE carcinoma.


Assuntos
Células Neuroendócrinas/patologia , Tumores Neuroendócrinos/patologia , Neoplasias da Próstata/patologia , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Carcinoma de Células Pequenas/genética , Carcinoma de Células Pequenas/metabolismo , Carcinoma de Células Pequenas/patologia , Diferenciação Celular/fisiologia , Estudos de Coortes , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Células Neuroendócrinas/metabolismo , Tumores Neuroendócrinos/genética , Tumores Neuroendócrinos/metabolismo , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/biossíntese , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Proteínas de Ligação a Retinoblastoma/genética , Proteínas de Ligação a Retinoblastoma/metabolismo , Estudos Retrospectivos , Transdução de Sinais , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
10.
PLoS Genet ; 16(7): e1008933, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32692737

RESUMO

Structure-specific endonucleases (SSEs) play key roles in DNA replication, recombination, and repair. SSEs must be tightly regulated to ensure genome stability but their regulatory mechanisms remain incompletely understood. Here, we show that in the fission yeast Schizosaccharomyces pombe, the activities of two SSEs, Dna2 and Rad16 (ortholog of human XPF), are temporally controlled during the cell cycle by the CRL4Cdt2 ubiquitin ligase. CRL4Cdt2 targets Pxd1, an inhibitor of Dna2 and an activator of Rad16, for degradation in S phase. The ubiquitination and degradation of Pxd1 is dependent on CRL4Cdt2, PCNA, and a PCNA-binding degron motif on Pxd1. CRL4Cdt2-mediated Pxd1 degradation prevents Pxd1 from interfering with the normal S-phase functions of Dna2. Moreover, Pxd1 degradation leads to a reduction of Rad16 nuclease activity in S phase, and restrains Rad16-mediated single-strand annealing, a hazardous pathway of repairing double-strand breaks. These results demonstrate a new role of the CRL4Cdt2 ubiquitin ligase in genome stability maintenance and shed new light on how SSE activities are regulated during the cell cycle.


Assuntos
Proteínas de Ligação a DNA/genética , Endonucleases Flap/genética , Proteínas Nucleares/genética , Proteínas de Schizosaccharomyces pombe/genética , Reparo do DNA/genética , Replicação do DNA/genética , Instabilidade Genômica/genética , Humanos , Fase S/genética , Schizosaccharomyces/genética , Ubiquitina/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação/genética
11.
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
12.
Molecules ; 25(12)2020 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-32604797

RESUMO

Viruses can be spread from one person to another; therefore, they may cause disorders in many people, sometimes leading to epidemics and even pandemics. New, previously unstudied viruses and some specific mutant or recombinant variants of known viruses constantly appear. An example is a variant of coronaviruses (CoV) causing severe acute respiratory syndrome (SARS), named SARS-CoV-2. Some antiviral drugs, such as remdesivir as well as antiretroviral drugs including darunavir, lopinavir, and ritonavir are suggested to be effective in treating disorders caused by SARS-CoV-2. There are data on the utilization of antiretroviral drugs against SARS-CoV-2. Since there are many studies aimed at the identification of the molecular mechanisms of human immunodeficiency virus type 1 (HIV-1) infection and the development of novel therapeutic approaches against HIV-1, we used HIV-1 for our case study to identify possible molecular pathways shared by SARS-CoV-2 and HIV-1. We applied a text and data mining workflow and identified a list of 46 targets, which can be essential for the development of infections caused by SARS-CoV-2 and HIV-1. We show that SARS-CoV-2 and HIV-1 share some molecular pathways involved in inflammation, immune response, cell cycle regulation.


Assuntos
Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/metabolismo , Mineração de Dados/métodos , Infecções por HIV/epidemiologia , Infecções por HIV/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/metabolismo , Anti-Inflamatórios/uso terapêutico , Antígenos de Diferenciação/genética , Antígenos de Diferenciação/imunologia , Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/imunologia , Betacoronavirus/patogenicidade , Proteínas do Sistema Complemento/genética , Proteínas do Sistema Complemento/imunologia , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Bases de Dados Genéticas , Regulação da Expressão Gênica , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , HIV-1/efeitos dos fármacos , HIV-1/imunologia , HIV-1/patogenicidade , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Interações Hospedeiro-Patógeno/genética , Humanos , Imunidade Inata/efeitos dos fármacos , Fatores Imunológicos/uso terapêutico , Inflamação , Interferons/genética , Interferons/imunologia , Interleucinas/genética , Interleucinas/imunologia , Redes e Vias Metabólicas/efeitos dos fármacos , Redes e Vias Metabólicas/genética , Redes e Vias Metabólicas/imunologia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/imunologia , Proteínas Repressoras/genética , Proteínas Repressoras/imunologia , Transdução de Sinais , Receptores Toll-Like/genética , Receptores Toll-Like/imunologia , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/imunologia
13.
Plant Mol Biol ; 104(1-2): 81-95, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32621166

RESUMO

KEY MESSAGE: Genome-wide identification of WD40-like genes reveals a duplication of COP1-like genes, one of the key players involved in regulation of flowering time and photomorphogenesis, with strong functional diversification in Rosaceae. WD40 proteins play crucial roles in a broad spectrum of developmental and physiological processes. Here, we conducted a systematic characterization of this family of genes in Rosa chinensis 'Old Blush' (OB), a founder genotype for modern rose domestication. We identified 187 rose WD40 genes and classified them into 5 clusters and 15 subfamilies with 11 of RcWD40s presumably generated via tandem duplication. We found RcWD40 genes were expressed differentially following stages of vegetative and reproductive development. We detected a duplication of CONSTITUTIVE PHOTOMORPHOGENIC1-like genes in rose (RcCOP1 and RcCOP1L) and other Rosaceae plants. Featuring a distinct expression pattern and a different profile of cis-regulatory-elements in the transcriptional regulatory regions, RcCOP1 seemed being evolutionarily conserved while RcCOP1L did not dimerize with RcHY5 and RcSPA4. Our data thus reveals a functional diversification of COP1-like genes in Rosacaeae plants, and provides a valuable resource to explore the potential function and evolution of WD40-like genes in Rosaceae plants.


Assuntos
Genes de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Rosaceae/genética , Rosaceae/metabolismo , Ubiquitina-Proteína Ligases/genética , Sequência de Aminoácidos , Arabidopsis/genética , Arabidopsis/metabolismo , Cromossomos de Plantas/genética , Domesticação , Duplicação Gênica , Regulação da Expressão Gênica de Plantas , Genoma de Planta , Filogenia , Plantas Geneticamente Modificadas , Rosa/genética , Rosa/metabolismo , Ubiquitina-Proteína Ligases/metabolismo
14.
Dev Genes Evol ; 230(4): 279-294, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32623522

RESUMO

Genome studies have uncovered many examples of essential gene loss, raising the question of how ancient genes transition from essentiality to dispensability. We explored this process for the deeply conserved E3 ubiquitin ligase Murine double minute (Mdm), which is lacking in Drosophila despite the conservation of its main regulatory target, the cellular stress response gene p53. Conducting gene expression and knockdown experiments in the red flour beetle Tribolium castaneum, we found evidence that Mdm has remained essential in insects where it is present. Using bioinformatics approaches, we confirm the absence of the Mdm gene family in Drosophila, mapping its loss to the stem lineage of schizophoran Diptera and Pipunculidae (big-headed flies), about 95-85 million years ago. Intriguingly, this gene loss event was preceded by the de novo origin of the gene Companion of reaper (Corp), a novel p53 regulatory factor that is characterized by functional similarities to vertebrate Mdm2 despite lacking E3 ubiquitin ligase protein domains. Speaking against a 1:1 compensatory gene gain/loss scenario, however, we found that hoverflies (Syrphidae) and pointed-wing flies (Lonchopteridae) possess both Mdm and Corp. This implies that the two p53 regulators have been coexisting for ~ 150 million years in select dipteran clades and for at least 50 million years in the lineage to Schizophora and Pipunculidae. Given these extensive time spans of Mdm/Corp coexistence, we speculate that the loss of Mdm in the lineage to Drosophila involved further acquisitions of compensatory gene activities besides the emergence of Corp. Combined with the previously noted reduction of an ancestral P53 contact domain in the Mdm homologs of crustaceans and insects, we conclude that the loss of the ancient Mdm gene family in flies was the outcome of incremental functional regression over long macroevolutionary time scales.


Assuntos
Proteínas de Drosophila/genética , Drosophila/genética , Genes Essenciais/genética , Proteínas Proto-Oncogênicas c-mdm2/genética , Tribolium/genética , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína Ligases/genética , Animais , Evolução Molecular , Técnicas de Silenciamento de Genes , Genômica , Filogenia , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Tribolium/embriologia , Proteína Supressora de Tumor p53/genética
15.
Proc Natl Acad Sci U S A ; 117(32): 19190-19200, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32723828

RESUMO

The 26S proteasome, a self-compartmentalized protease complex, plays a crucial role in protein quality control. Multiple levels of regulatory systems modulate proteasomal activity for substrate hydrolysis. However, the destruction mechanism of mammalian proteasomes is poorly understood. We found that inhibited proteasomes are sequestered into the insoluble aggresome via HDAC6- and dynein-mediated transport. These proteasomes colocalized with the autophagic receptor SQSTM1 and cleared through selective macroautophagy, linking aggresomal segregation to autophagic degradation. This proteaphagic pathway was counterbalanced with the recovery of proteasomal activity and was critical for reducing cellular proteasomal stress. Changes in associated proteins and polyubiquitylation on inhibited 26S proteasomes participated in the targeting mechanism to the aggresome and autophagosome. The STUB1 E3 Ub ligase specifically ubiquitylated purified human proteasomes in vitro, mainly via Lys63-linked chains. Genetic and chemical inhibition of STUB1 activity significantly impaired proteasome processing and reduced resistance to proteasomal stress. These data demonstrate that aggresomal sequestration is the crucial upstream event for proteasome quality control and overall protein homeostasis in mammals.


Assuntos
Macroautofagia , Organelas/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Células A549 , Humanos , Organelas/genética , Complexo de Endopeptidases do Proteassoma/genética , Proteína Sequestossoma-1/genética , Proteína Sequestossoma-1/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
16.
Nucleic Acids Res ; 48(14): 7728-7747, 2020 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-32609811

RESUMO

UHRF1 is an important epigenetic regulator associated with apoptosis and tumour development. It is a multidomain protein that integrates readout of different histone modification states and DNA methylation with enzymatic histone ubiquitylation activity. Emerging evidence indicates that the chromatin-binding and enzymatic modules of UHRF1 do not act in isolation but interplay in a coordinated and regulated manner. Here, we compared two splicing variants (V1, V2) of murine UHRF1 (mUHRF1) with human UHRF1 (hUHRF1). We show that insertion of nine amino acids in a linker region connecting the different TTD and PHD histone modification-binding domains causes distinct H3K9me3-binding behaviour of mUHRF1 V1. Structural analysis suggests that in mUHRF1 V1, in contrast to V2 and hUHRF1, the linker is anchored in a surface groove of the TTD domain, resulting in creation of a coupled TTD-PHD module. This establishes multivalent, synergistic H3-tail binding causing distinct cellular localization and enhanced H3K9me3-nucleosome ubiquitylation activity. In contrast to hUHRF1, H3K9me3-binding of the murine proteins is not allosterically regulated by phosphatidylinositol 5-phosphate that interacts with a separate less-conserved polybasic linker region of the protein. Our results highlight the importance of flexible linkers in regulating multidomain chromatin binding proteins and point to divergent evolution of their regulation.


Assuntos
Processamento Alternativo , Proteínas Estimuladoras de Ligação a CCAAT/química , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Histonas/metabolismo , Ubiquitina-Proteína Ligases/química , Ubiquitina-Proteína Ligases/metabolismo , Regulação Alostérica , Animais , Proteínas Estimuladoras de Ligação a CCAAT/genética , Linhagem Celular , Núcleo Celular/metabolismo , Cromatina/metabolismo , Código das Histonas , Humanos , Camundongos , Ligação Proteica , Domínio Tudor , Ubiquitina-Proteína Ligases/genética
17.
Mol Cell ; 79(4): 588-602.e6, 2020 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-32615089

RESUMO

The ribosome-associated protein quality control (RQC) system that resolves stalled translation events is activated when ribosomes collide and form disome, trisome, or higher-order complexes. However, it is unclear whether this system distinguishes collision complexes formed on defective mRNAs from those with functional roles on endogenous transcripts. Here, we performed disome and trisome footprint profiling in yeast and found collisions were enriched on diverse sequence motifs known to slow translation. When 60S recycling was inhibited, disomes accumulated at stop codons and could move into the 3' UTR to reinitiate translation. The ubiquitin ligase and RQC factor Hel2/ZNF598 generally recognized collisions but did not induce degradation of endogenous transcripts. However, loss of Hel2 triggered the integrated stress response, via phosphorylation of eIF2α, thus linking these pathways. Our results suggest that Hel2 has a role in sensing ribosome collisions on endogenous mRNAs, and such events may be important for cellular homeostasis.


Assuntos
Pegada de DNA/métodos , Genoma Fúngico , Ribossomos/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Ubiquitina-Proteína Ligases/metabolismo , Regiões 3' não Traduzidas , Anisomicina/farmacologia , Códon de Terminação , Fator de Iniciação 2 em Eucariotos/genética , Fator de Iniciação 2 em Eucariotos/metabolismo , Mutação , Fosforilação , Estabilidade de RNA , Subunidades Ribossômicas Maiores de Eucariotos/genética , Subunidades Ribossômicas Maiores de Eucariotos/metabolismo , Ribossomos/metabolismo , Saccharomyces cerevisiae/efeitos dos fármacos , Proteínas de Saccharomyces cerevisiae/genética , Ubiquitina-Proteína Ligases/genética
18.
Proc Natl Acad Sci U S A ; 117(28): 16567-16578, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32606244

RESUMO

Malaria infection induces complex and diverse immune responses. To elucidate the mechanisms underlying host-parasite interaction, we performed a genetic screen during early (24 h) Plasmodium yoelii infection in mice and identified a large number of interacting host and parasite genes/loci after transspecies expression quantitative trait locus (Ts-eQTL) analysis. We next investigated a host E3 ubiquitin ligase gene (March1) that was clustered with interferon (IFN)-stimulated genes (ISGs) based on the similarity of the genome-wide pattern of logarithm of the odds (LOD) scores (GPLS). March1 inhibits MAVS/STING/TRIF-induced type I IFN (IFN-I) signaling in vitro and in vivo. However, in malaria-infected hosts, deficiency of March1 reduces IFN-I production by activating inhibitors such as SOCS1, USP18, and TRIM24 and by altering immune cell populations. March1 deficiency increases CD86+DC (dendritic cell) populations and levels of IFN-γ and interleukin 10 (IL-10) at day 4 post infection, leading to improved host survival. T cell depletion reduces IFN-γ level and reverse the protective effects of March1 deficiency, which can also be achieved by antibody neutralization of IFN-γ. This study reveals functions of MARCH1 (membrane-associated ring-CH-type finger 1) in innate immune responses and provides potential avenues for activating antimalaria immunity and enhancing vaccine efficacy.


Assuntos
Malária/imunologia , Plasmodium yoelii/fisiologia , Linfócitos T/imunologia , Ubiquitina-Proteína Ligases/imunologia , Animais , Modelos Animais de Doenças , Feminino , Interações Hospedeiro-Parasita , Humanos , Imunidade Inata , Interferon Tipo I/genética , Interferon Tipo I/imunologia , Interferon gama/genética , Interferon gama/imunologia , Interleucina-10/genética , Interleucina-10/imunologia , Malária/enzimologia , Malária/genética , Malária/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Plasmodium yoelii/imunologia , Ubiquitina-Proteína Ligases/genética
19.
Nat Commun ; 11(1): 3615, 2020 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-32680986

RESUMO

Failure to preserve the integrity of the genome is a hallmark of cancer. Recent studies have revealed that loss of the capacity to repair DNA breaks via homologous recombination (HR) results in a mutational profile termed BRCAness. The enzymatic activity that repairs HR substrates in BRCA-deficient conditions to produce this profile is currently unknown. We here show that the mutational landscape of BRCA1 deficiency in C. elegans closely resembles that of BRCA1-deficient tumours. We identify polymerase theta-mediated end-joining (TMEJ) to be responsible: knocking out polq-1 suppresses the accumulation of deletions and tandem duplications in brc-1 and brd-1 animals. We find no additional back-up repair in HR and TMEJ compromised animals; non-homologous end-joining does not affect BRCAness. The notion that TMEJ acts as an alternative to HR, promoting the genome alteration of HR-deficient cells, supports the idea that polymerase theta is a promising therapeutic target for HR-deficient tumours.


Assuntos
Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Reparo do DNA por Junção de Extremidades , DNA Polimerase Dirigida por DNA/metabolismo , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Animais , Animais Geneticamente Modificados , Análise Mutacional de DNA , DNA Polimerase Dirigida por DNA/genética , Técnicas de Inativação de Genes , Mutação
20.
PLoS One ; 15(7): e0235925, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32639967

RESUMO

Angelman syndrome (AS) is a rare neurodevelopmental disorder characterized by speech impairment, intellectual disability, ataxia, and epilepsy. AS is caused by mutations in the maternal copy of UBE3A located on chromosome 15q11-13. UBE3A codes for E6AP (E6 Associated Protein), a prominent member of the HECT (Homologous to E6AP C-Terminus) E3 ubiquitin ligase family. E6AP catalyzes the posttranslational attachment of ubiquitin via its HECT domain onto various intracellular target proteins to regulate DNA repair and cell cycle progression. The HECT domain consists of an N-lobe, required for E2~ubiquitin recruitment, while the C-lobe contains the conserved catalytic cysteine required for ubiquitin transfer. Previous genetic studies of AS patients have identified point mutations in UBE3A that result in amino acid substitutions or premature termination during translation. An AS transversion mutation (codon change from ATA to AAA) within the region of the gene that codes for the catalytic HECT domain of E6AP has been annotated (I827K), but the molecular basis for this loss of function substitution remained elusive. Here, we demonstrate that the I827K substitution destabilizes the 3D fold causing protein aggregation of the C-terminal lobe of E6AP using a combination of spectropolarimetry and nuclear magnetic resonance (NMR) spectroscopy. Our fluorescent ubiquitin activity assays with E6AP-I827K show decreased ubiquitin thiolester formation and ubiquitin discharge. Using 3D models in combination with our biochemical and biophysical results, we rationalize why the I827K disrupts E6AP-dependent ubiquitylation. This work provides new insight into the E6AP mechanism and how its malfunction can be linked to the AS phenotype.


Assuntos
Síndrome de Angelman/genética , Ubiquitina-Proteína Ligases/metabolismo , Síndrome de Angelman/patologia , Biocatálise , Dicroísmo Circular , Humanos , Mutagênese Sítio-Dirigida , Ressonância Magnética Nuclear Biomolecular , Domínios Proteicos , Estabilidade Proteica , Estrutura Secundária de Proteína , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/química , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
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