Epstein-Barr virus （EBV）-encoded latent membrane protein 1 （LMPI） has been known to have oncogenic properties during latent infection in nasopharyngeal carcinoma （NPC）. Our studies focused on the role of LMP1 in...Epstein-Barr virus （EBV）-encoded latent membrane protein 1 （LMPI） has been known to have oncogenic properties during latent infection in nasopharyngeal carcinoma （NPC）. Our studies focused on the role of LMP1 in NPC, and showed that LMP1 triggers the NF-κB, AP-1 and STAT signaling pathways. Strikingly, LMP1 was found to mediate the formation of a new heterodimer between c-Jun and JunB. Also, we have identified JAK/STAT and PI-PLC-PKC activation triggered by LMP1 through upregulating the expression of JAK3 and enhancing the phosphorylation of STATo The constitutive activation of these signaling cascades explains LMP1＇s ability to induce such a diverse array of morphological and phenotypic effects in cells and provides insight into how LMP1 may induce cell transformation, in which multihit targeted genes in the downstream play an essential role. All signaling cascades triggered by LMP1 ultimately lead to the disruption of the cell cycle： the acceleration of G1/S phase and the arrest of G2/M phase. We also found that LMP1 induced the expression of hTERT and promoted cell immortalization. Importantly, by intervening physical intracellular signal transduction pathways and disturbing the progression of the cell cycle, LMP1, an important oncoprotein encoded by EBV, is thought to be a key modulator in the pathogenesis of NPC. Interfering LMP1 signaling could be a promising strategy to target the malignant phenotype of NPC. Cellular ＆ Molecular Immunology.展开更多
Background:Natural killer/T-cell lymphoma(NKTCL)is a highly aggressive non-Hodgkin lymphoma often resistant to chemotherapy.Serum level of soluble IL-2 receptorα(IL-2Rα)is elevated in NKTCL patients and correlates s...Background:Natural killer/T-cell lymphoma(NKTCL)is a highly aggressive non-Hodgkin lymphoma often resistant to chemotherapy.Serum level of soluble IL-2 receptorα(IL-2Rα)is elevated in NKTCL patients and correlates signifi-cantly with treatment response and survival.In the current study we examined the potential role of IL-2Rαby over-expressing IL-2Rαin representative cell lines.Methods:Levels of IL-2Rαwere evaluated in the human natural killer cell line NK-92 and the NKTCL cell line SNK-6.Lentiviral vectors were used to express latent membrane protein 1(LMP1)in NK-92 cells,and IL-2Rαin both NK-92 and SNK-6 cells.The biological effects of these genes on proliferation,apoptosis,cell cycle distribution,and chemosensitiv-ity were analyzed.Results:Expression of IL-2Rαwas significantly higher in SNK-6 cells than in NK-92 cells.Expressing LMP1 in NK-92 cells remarkably up-regulated IL-2Rαlevels,whereas selective inhibitorss of the proteins in the MAPK/NF-κB pathway significantly down-regulated IL-2Rα.IL-2Rαoverexpression in SNK-6 cells promoted cell proliferation by altering cell cycle distribution,and induced resistance to gemcitabine,doxorubicin,and asparaginase.These effects were reversed by an anti-IL-2Rαantibody.Conclusions:Our results suggest that LMP1 activates the MAPK/NF-κB pathway in NKTCL cells,up-regulating IL-2Rαexpression.IL-2Rαoverexpression promotes growth and chemoresistance in NKTCL,making this interleukin receptor a potential therapeutic target.展开更多
Latent membrane protein 1 （LMP1）, an important protein encoded by Epstein Barr virus （EBV）, has been implied to link with the pathogenesis of nasopharyngeal carcinoma （NPC）. Its dual effects of increasing cell p...Latent membrane protein 1 （LMP1）, an important protein encoded by Epstein Barr virus （EBV）, has been implied to link with the pathogenesis of nasopharyngeal carcinoma （NPC）. Its dual effects of increasing cell proliferation and inhibiting cell apoptosis have been confirmed. In this study, we showed that the expression of Survivin and CDK4 protein in CNE-LMP1, a LMP1 positive NPC epithelial cell line, is higher than in LMP1 negative NPC epithelial cell line- CNE1, and the expression is LMP1 dosage-dependent. Although it was reported that Survivin specifically expressed in cell cycle G2/M phase, our studies suggested that LMP1 could promote the expression of Survivin in G0/G1, S and G2/ M phase. It also showed that Survivin and CDK4 could be accumulated more in the nuclei triggered by LMP1. More interestingly, Survivin and CDK4 could form a protein complex in the nuclei of CNE-LMP1 rather than in that of CNE1, which demonstrated that the interaction between these two proteins could be promoted by LMPI. These results strongly suggested that the role of LMP1 in the regulation of Survivin and CDK4 may also shed some light on the mechanism research of LMP1 in NPC.展开更多
Back ground: Non-Hodgkin’s lymphoma (NHL) with its different subtypes is strongly related to Epstein Bar virus (EBV) infection mainly Burkitt’s lymphoma in Africa. Studies proved the role of EBV in tumor-genesis and...Back ground: Non-Hodgkin’s lymphoma (NHL) with its different subtypes is strongly related to Epstein Bar virus (EBV) infection mainly Burkitt’s lymphoma in Africa. Studies proved the role of EBV in tumor-genesis and linked it to prognosis and therapy of patients. Objectives: To determine the frequency of EBV in non-Hodgkin lymphomas using EBV latent membrane protein 1 (EBV-LMP1) immunohistochemical stain. Methods: This cross-sectional study was conducted at radio-isotope centre of Khartoum (2012-2014). A total of 75 cases of non-Hodgkin lymphoma paraffin embedded sections were stained for EBV LMP1 antibody. Data were analyzed by SPSS 16 and statistical cross linking of the results of immune staining with other data was done. Results: Out of 75 patients of non Hodgkin’s lymphoma (74.7%) were males. EBV-LMP1 Immune-staining was positive in (17.3%) with predominance of Burkitt’s lymphoma (33.3%), followed by diffuse large B cell lymphoma (17.9%). Conclusion: Burkitt’s lymphoma expressed the highest percentage of non-Hodgkin’s lymphoma positive cases (46.2%) out of the total (17.3%) positive cases. Different methods need to be used in studying Burkitt’s lymphoma expression of EBV and its effects on the treatment and prognosis of cases.展开更多
Background:Immunotherapy has revolutionized the therapeutical regimen for nasopharyngeal carcinoma(NPC),yet its response rate remains insufficient.Programmed death-ligand 1(PD-L1)on small extracellular vesicles(sEVs)m...Background:Immunotherapy has revolutionized the therapeutical regimen for nasopharyngeal carcinoma(NPC),yet its response rate remains insufficient.Programmed death-ligand 1(PD-L1)on small extracellular vesicles(sEVs)mediates local and peripheral immunosuppression in tumors,and the mechanism of PD-L1 loading into these vesicles is garnering increasing attention.Latent membrane protein 1(LMP1),a key viral oncoprotein expressed in Epstein-Barr virus(EBV)-positive NPC,contributes to remodeling the tumor microenvironment.However,the precise mechanisms by which LMP1 modulates tumor immunity in NPC remain unclear.Here,we aimed to investigate the roles and regulatory mechanisms of LMP1 and sEV PD-L1 in NPC immune evasion.Methods:We analyzed the impact of LMP1 on tumor-infiltrating lymphocyte abundance in NPC tissues and humanized tumor-bearing mouse models using multiplex immunofluorescence(mIF)and flow cytometry,respectively.Transmission electron microscopy and nanoparticle tracking analysis were employed to characterize sEVs.Immunoprecipitation-mass spectrometry was utilized to identify proteins interacting with LMP1.The regulatory effects of sEVs on tumor microenvironment were assessed by monitoring CD8^(+)T cell proliferation and interferon-γ(IFN-γ)expression via flow cytometry.Furthermore,the expression patterns of LMP1 and downstream regulators in NPC were analyzed using mIF and survival analysis.Results:High LMP1 expression in NPC patient specimens and mouse models was associated with restricted infiltration of CD8^(+)T cells.Additionally,LMP1 promoted sEV PD-L1 secretion,leading to inhibition of CD8^(+)T cell viability and IFN-γ expression in vitro.Mechanistically,LMP1 recruited apoptosis-linked gene 2-interacting protein X(ALIX)through its intracellular domain and bound PD-L1 through its transmembrane domain,thereby facilitating the loading of PDL1 into ALIX-dependent sEVs.Disruption of ALIX diminished LMP1-induced sEV PD-L1 secretion and enhanced the anti-tumor immunity of CD8^(+)T cells both in vitro and in vivo.Moreover,increased expression levels of LMP1 and ALIXwere positively correlated with enhanced immunosuppressive features and worse prognostic outcomes in NPC patients.Conclusion:Our findings uncovered the mechanism by which LMP1 interacts with ALIX and PD-L1 to form a trimolecular complex,facilitating PD-L1 loading into ALIX-dependent sEV secretion pathway,ultimately inhibiting the antitumor immune response in NPC.This highlights a novel target and prognostic marker for NPC immunotherapy.展开更多
The AKT/mTOR and NF-κB signalings are crucial pathways activated in cancers including nasopharyngeal carcinoma(NPC), which is prevalent in southern China and closely related to Epstein-Barr virus(EBV) infection.How t...The AKT/mTOR and NF-κB signalings are crucial pathways activated in cancers including nasopharyngeal carcinoma(NPC), which is prevalent in southern China and closely related to Epstein-Barr virus(EBV) infection.How these master pathways are persistently activated in EBV-associated NPC remains to be investigated. Here we demonstrated that EBV-encoded latent membrane protein 1(LMP1) promoted cyclophilin A(CYPA) expression through the activation of NF-κB. The depletion of CYPA suppressed cell proliferation and facilitated apoptosis.CYPA was able to bind to AKT1, thus activating AKT/mTOR/NF-κB signaling cascade. Moreover, the use of mTOR inhibitor, rapamycin, subverted the activation of the positive feedback loop, NF-κB/CYPA/AKT/mTOR. It is reasonable that LMP1 expression derived from initial viral infection is enough to assure the constant potentiation of AKT/mTOR and NF-κB signalings. This may partly explain the fact that EBV serves as a tumor-promoting factor with minimal expression of the viral oncoprotein LMP1 in malignancies. Our findings provide new insight into the understanding of causative role of EBV in tumorigenicity during latent infection.展开更多
基金grants from State Key Basic Research and Development Plan （973） of the Ministry of Science and Technology of China （2004CB518703） National High Technology Research and Development Program （863） of China （N0.2006AA02Z481）+1 种基金 National Science Fund for Distinguished Young Scholars 0verseas （30428008） Hong Kong Cooperation Fund Project of National Natural Science Foundation of China （30418004）.
文摘Epstein-Barr virus （EBV）-encoded latent membrane protein 1 （LMPI） has been known to have oncogenic properties during latent infection in nasopharyngeal carcinoma （NPC）. Our studies focused on the role of LMP1 in NPC, and showed that LMP1 triggers the NF-κB, AP-1 and STAT signaling pathways. Strikingly, LMP1 was found to mediate the formation of a new heterodimer between c-Jun and JunB. Also, we have identified JAK/STAT and PI-PLC-PKC activation triggered by LMP1 through upregulating the expression of JAK3 and enhancing the phosphorylation of STATo The constitutive activation of these signaling cascades explains LMP1＇s ability to induce such a diverse array of morphological and phenotypic effects in cells and provides insight into how LMP1 may induce cell transformation, in which multihit targeted genes in the downstream play an essential role. All signaling cascades triggered by LMP1 ultimately lead to the disruption of the cell cycle： the acceleration of G1/S phase and the arrest of G2/M phase. We also found that LMP1 induced the expression of hTERT and promoted cell immortalization. Importantly, by intervening physical intracellular signal transduction pathways and disturbing the progression of the cell cycle, LMP1, an important oncoprotein encoded by EBV, is thought to be a key modulator in the pathogenesis of NPC. Interfering LMP1 signaling could be a promising strategy to target the malignant phenotype of NPC. Cellular ＆ Molecular Immunology.
基金the National Natural Science Foundation of China(81400159,81873450,81700196,81672686)Pearl River Nova Program of Guangzhou(201710010161)+1 种基金Sister Institution Net-work Fund of the MD Anderson Cancer Center(to Qingqing Cai)the Fundamental Research Funds for the Central Universities(17ykpy77).
文摘Background:Natural killer/T-cell lymphoma(NKTCL)is a highly aggressive non-Hodgkin lymphoma often resistant to chemotherapy.Serum level of soluble IL-2 receptorα(IL-2Rα)is elevated in NKTCL patients and correlates signifi-cantly with treatment response and survival.In the current study we examined the potential role of IL-2Rαby over-expressing IL-2Rαin representative cell lines.Methods:Levels of IL-2Rαwere evaluated in the human natural killer cell line NK-92 and the NKTCL cell line SNK-6.Lentiviral vectors were used to express latent membrane protein 1(LMP1)in NK-92 cells,and IL-2Rαin both NK-92 and SNK-6 cells.The biological effects of these genes on proliferation,apoptosis,cell cycle distribution,and chemosensitiv-ity were analyzed.Results:Expression of IL-2Rαwas significantly higher in SNK-6 cells than in NK-92 cells.Expressing LMP1 in NK-92 cells remarkably up-regulated IL-2Rαlevels,whereas selective inhibitorss of the proteins in the MAPK/NF-κB pathway significantly down-regulated IL-2Rα.IL-2Rαoverexpression in SNK-6 cells promoted cell proliferation by altering cell cycle distribution,and induced resistance to gemcitabine,doxorubicin,and asparaginase.These effects were reversed by an anti-IL-2Rαantibody.Conclusions:Our results suggest that LMP1 activates the MAPK/NF-κB pathway in NKTCL cells,up-regulating IL-2Rαexpression.IL-2Rαoverexpression promotes growth and chemoresistance in NKTCL,making this interleukin receptor a potential therapeutic target.
基金National Nature Science Foundation for Distinguished Young Scholar of China (No.39525022)National Basic Research Program(No.2004CB518703) National Nature Science Foundation of China (No.30570085).
文摘Latent membrane protein 1 （LMP1）, an important protein encoded by Epstein Barr virus （EBV）, has been implied to link with the pathogenesis of nasopharyngeal carcinoma （NPC）. Its dual effects of increasing cell proliferation and inhibiting cell apoptosis have been confirmed. In this study, we showed that the expression of Survivin and CDK4 protein in CNE-LMP1, a LMP1 positive NPC epithelial cell line, is higher than in LMP1 negative NPC epithelial cell line- CNE1, and the expression is LMP1 dosage-dependent. Although it was reported that Survivin specifically expressed in cell cycle G2/M phase, our studies suggested that LMP1 could promote the expression of Survivin in G0/G1, S and G2/ M phase. It also showed that Survivin and CDK4 could be accumulated more in the nuclei triggered by LMP1. More interestingly, Survivin and CDK4 could form a protein complex in the nuclei of CNE-LMP1 rather than in that of CNE1, which demonstrated that the interaction between these two proteins could be promoted by LMPI. These results strongly suggested that the role of LMP1 in the regulation of Survivin and CDK4 may also shed some light on the mechanism research of LMP1 in NPC.
文摘Back ground: Non-Hodgkin’s lymphoma (NHL) with its different subtypes is strongly related to Epstein Bar virus (EBV) infection mainly Burkitt’s lymphoma in Africa. Studies proved the role of EBV in tumor-genesis and linked it to prognosis and therapy of patients. Objectives: To determine the frequency of EBV in non-Hodgkin lymphomas using EBV latent membrane protein 1 (EBV-LMP1) immunohistochemical stain. Methods: This cross-sectional study was conducted at radio-isotope centre of Khartoum (2012-2014). A total of 75 cases of non-Hodgkin lymphoma paraffin embedded sections were stained for EBV LMP1 antibody. Data were analyzed by SPSS 16 and statistical cross linking of the results of immune staining with other data was done. Results: Out of 75 patients of non Hodgkin’s lymphoma (74.7%) were males. EBV-LMP1 Immune-staining was positive in (17.3%) with predominance of Burkitt’s lymphoma (33.3%), followed by diffuse large B cell lymphoma (17.9%). Conclusion: Burkitt’s lymphoma expressed the highest percentage of non-Hodgkin’s lymphoma positive cases (46.2%) out of the total (17.3%) positive cases. Different methods need to be used in studying Burkitt’s lymphoma expression of EBV and its effects on the treatment and prognosis of cases.
基金Health Commission of Hubei Province Medical Leading Talent Project,and Translational Medicine and Interdisciplinary Research Joint Fund of Zhongnan Hospital of Wuhan University,Grant/Award Numbers:ZNJC201922,ZNJC202007Key Research&Development Project of Hubei Province,Grant/Award Number:2020BCA069+1 种基金National Natural Science Foundation of China,Grant/Award Numbers:81803065,81972852Research Projects of Biomedical Center of Hubei Cancer Hospital,Grant/Award Number:2022SWZX12。
文摘Background:Immunotherapy has revolutionized the therapeutical regimen for nasopharyngeal carcinoma(NPC),yet its response rate remains insufficient.Programmed death-ligand 1(PD-L1)on small extracellular vesicles(sEVs)mediates local and peripheral immunosuppression in tumors,and the mechanism of PD-L1 loading into these vesicles is garnering increasing attention.Latent membrane protein 1(LMP1),a key viral oncoprotein expressed in Epstein-Barr virus(EBV)-positive NPC,contributes to remodeling the tumor microenvironment.However,the precise mechanisms by which LMP1 modulates tumor immunity in NPC remain unclear.Here,we aimed to investigate the roles and regulatory mechanisms of LMP1 and sEV PD-L1 in NPC immune evasion.Methods:We analyzed the impact of LMP1 on tumor-infiltrating lymphocyte abundance in NPC tissues and humanized tumor-bearing mouse models using multiplex immunofluorescence(mIF)and flow cytometry,respectively.Transmission electron microscopy and nanoparticle tracking analysis were employed to characterize sEVs.Immunoprecipitation-mass spectrometry was utilized to identify proteins interacting with LMP1.The regulatory effects of sEVs on tumor microenvironment were assessed by monitoring CD8^(+)T cell proliferation and interferon-γ(IFN-γ)expression via flow cytometry.Furthermore,the expression patterns of LMP1 and downstream regulators in NPC were analyzed using mIF and survival analysis.Results:High LMP1 expression in NPC patient specimens and mouse models was associated with restricted infiltration of CD8^(+)T cells.Additionally,LMP1 promoted sEV PD-L1 secretion,leading to inhibition of CD8^(+)T cell viability and IFN-γ expression in vitro.Mechanistically,LMP1 recruited apoptosis-linked gene 2-interacting protein X(ALIX)through its intracellular domain and bound PD-L1 through its transmembrane domain,thereby facilitating the loading of PDL1 into ALIX-dependent sEVs.Disruption of ALIX diminished LMP1-induced sEV PD-L1 secretion and enhanced the anti-tumor immunity of CD8^(+)T cells both in vitro and in vivo.Moreover,increased expression levels of LMP1 and ALIXwere positively correlated with enhanced immunosuppressive features and worse prognostic outcomes in NPC patients.Conclusion:Our findings uncovered the mechanism by which LMP1 interacts with ALIX and PD-L1 to form a trimolecular complex,facilitating PD-L1 loading into ALIX-dependent sEV secretion pathway,ultimately inhibiting the antitumor immune response in NPC.This highlights a novel target and prognostic marker for NPC immunotherapy.
基金supported by the Natural Science Foundations of China(81974427)Graduate Research and Innovation Projects of Central South University(2021zzts0931)
文摘The AKT/mTOR and NF-κB signalings are crucial pathways activated in cancers including nasopharyngeal carcinoma(NPC), which is prevalent in southern China and closely related to Epstein-Barr virus(EBV) infection.How these master pathways are persistently activated in EBV-associated NPC remains to be investigated. Here we demonstrated that EBV-encoded latent membrane protein 1(LMP1) promoted cyclophilin A(CYPA) expression through the activation of NF-κB. The depletion of CYPA suppressed cell proliferation and facilitated apoptosis.CYPA was able to bind to AKT1, thus activating AKT/mTOR/NF-κB signaling cascade. Moreover, the use of mTOR inhibitor, rapamycin, subverted the activation of the positive feedback loop, NF-κB/CYPA/AKT/mTOR. It is reasonable that LMP1 expression derived from initial viral infection is enough to assure the constant potentiation of AKT/mTOR and NF-κB signalings. This may partly explain the fact that EBV serves as a tumor-promoting factor with minimal expression of the viral oncoprotein LMP1 in malignancies. Our findings provide new insight into the understanding of causative role of EBV in tumorigenicity during latent infection.
