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1.
J Virol ; 96(14): e0051822, 2022 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-35862711

RESUMO

Protein-protein interactions (PPIs) are crucial for various biological processes. Epstein-Barr virus (EBV) proteins typically form complexes, regulating the replication and persistence of the viral genome in human cells. However, the role of EBV protein complexes under physiological conditions remains unclear. In this study, we performed comprehensive analyses of EBV PPIs in living cells using the NanoBiT system. We identified 195 PPIs, many of which have not previously been reported. Computational analyses of these PPIs revealed that BLRF2, which is only found in gammaherpesviruses, is a central protein in the structural network of EBV tegument proteins. To characterize the role of BLRF2, we generated two BLRF2 knockout EBV clones using CRISPR/Cas9. BLRF2 knockout significantly decreased the production of infectious virus particles, which was partially restored by exogenous BLRF2 expression. In addition, self-association of BLRF2 protein was found, and mutation of the residues crucial for the self-association affected stability of the protein. Our data imply that BLRF2 is a tegument network hub that plays important roles in progeny virion maturation. IMPORTANCE EBV remains a significant public health challenge, causing infectious mononucleosis and several cancer types. Therefore, the better understanding of the molecular mechanisms underlying EBV replication is of high clinical importance. As protein-protein interactions (PPIs) are major regulators of virus-associated pathogenesis, comprehensive analyses of PPIs are essential. Previous studies on PPIs in EBV or other herpesviruses have predominantly employed the yeast two-hybrid (Y2H) system, immunoprecipitation, and pulldown assays. Herein, using a novel luminescence-based method, we identified 195 PPIs, most of which have not previously been reported. Computational and functional analyses using knockout viruses revealed that BLRF2 plays a central role in the EBV life cycle, which makes it a valuable target for drug development.


Assuntos
Infecções por Vírus Epstein-Barr , Herpesvirus Humano 4 , Mapas de Interação de Proteínas , Proteínas Virais , Infecções por Vírus Epstein-Barr/virologia , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/fisiologia , Humanos , Proteínas Virais/genética , Replicação Viral
2.
Cancer Sci ; 112(12): 5088-5099, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34609775

RESUMO

Epstein-Barr virus-associated lymphoproliferative disease (EBV-LPD) is frequently fatal. Innate immunity plays a key role in protecting against pathogens and cancers. The stimulator of interferon genes (STING) is regarded as a key adaptor protein allowing DNA sensors recognizing exogenous cytosolic DNA to activate the type I interferon signaling cascade. In terms of EBV tumorigenicity, the role of STING remains elusive. Here we showed that treatment with the STING inhibitor, C-176, suppressed EBV-induced transformation in peripheral blood mononuclear cells. In an EBV-LPD mouse model, C-176 treatment also inhibited tumor formation and prolonged survival. Treatment with B cells alone did not affect EBV transformation, but suppression of EBV-induced transformation was observed in the presence of T cells. Even without direct B cell-T cell contact in a transwell system, the inhibitor reduced the transformation activity, indicating that intercellular communication by humoral factors was critical to prevent EBV-induced transformation. These findings suggest that inhibition of STING signaling pathway with C-176 could be a new therapeutic target of EBV-LPD.


Assuntos
Antineoplásicos/administração & dosagem , Transformação Celular Viral/efeitos dos fármacos , Infecções por Vírus Epstein-Barr/tratamento farmacológico , Linfoma de Células B/prevenção & controle , Proteínas de Membrana/antagonistas & inibidores , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Infecções por Vírus Epstein-Barr/imunologia , Células HEK293 , Herpesvirus Humano 4 , Humanos , Células Jurkat , Linfoma de Células B/imunologia , Linfoma de Células B/virologia , Camundongos , Análise de Sobrevida , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto
3.
Am J Surg Pathol ; 44(9): 1173-1183, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32349049

RESUMO

Epstein-Barr virus (EBV)-positive diffuse large B-cell lymphoproliferation encompasses a broad range of clinicopathologic findings, including specific subtypes, for example, EBV mucocutaneous ulcer. Here we reassessed 36 cases of primary EBV diffuse large B-cell lymphomas (16 men and 20 women; median age, 69.5 y; range, 35 to 84 y), including 8 immunosuppressed patients (Lugano stage II-IV; median age, 74 y), 7 nonimmunosuppressed patients with stage I disease (median age, 69 y), and 21 nonimmunosuppressed patients with stage II-IV disease (median age, 69 y). All immunosuppressed patients exhibited iatrogenic immunodeficiency and an ulcerative appearance, with ulcer sites including the stomach (1 patient), small intestine (6 patients), and rectum (1 patient). Four patients were in the setting of treated lymphoma-associated immunosuppression. Immunosuppressed patients had higher incidences of intestinal involvement (P=0.001) and perforation (n=2) compared with advanced stage nonimmunosuppressed patients. Among nonimmunosuppressed stage I patients, lesions were restricted to the stomach, none showed multiple lesions or elevated serum lactate dehydrogenase, and the overall survival curve plateaued, although it was not statistically significant (P=0.0581). One nonimmunosuppressed stage I patient with a polypoid lesion exhibited spontaneous regression within 2 months after diagnosis, while another with bulky disease pursued an aggressive clinical course. Nonimmunosuppressed stage I cases without bulky masses may be considered EBV mucocutaneous ulcer with local progression. Our results demonstrated that primary EBV gastrointestinal diffuse large B-cell lymphoma could be delineated into 3 groups based on immune status and clinical stage, revealing distinguishing features useful as a pragmatic guide for diagnostic and therapeutic approaches.


Assuntos
Infecções por Vírus Epstein-Barr/virologia , Neoplasias Gastrointestinais/virologia , Herpesvirus Humano 4/genética , Hospedeiro Imunocomprometido , Imunossupressores/efeitos adversos , Linfoma Difuso de Grandes Células B/virologia , RNA Viral/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Progressão da Doença , Infecções por Vírus Epstein-Barr/imunologia , Infecções por Vírus Epstein-Barr/patologia , Infecções por Vírus Epstein-Barr/terapia , Feminino , Neoplasias Gastrointestinais/imunologia , Neoplasias Gastrointestinais/patologia , Neoplasias Gastrointestinais/terapia , Herpesvirus Humano 4/imunologia , Humanos , Linfoma Difuso de Grandes Células B/imunologia , Linfoma Difuso de Grandes Células B/patologia , Linfoma Difuso de Grandes Células B/terapia , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Valor Preditivo dos Testes , Intervalo Livre de Progressão , Indução de Remissão , Estudos Retrospectivos , Fatores de Risco
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