A characteristic common to herpesviruses is the ability to establish a latent infection in the hosts, a transcriptionally active region has detected during latency as well as a set of RNA that are known as Latency Ass...A characteristic common to herpesviruses is the ability to establish a latent infection in the hosts, a transcriptionally active region has detected during latency as well as a set of RNA that are known as Latency Associated Transcripts (LATs), their functions have been clarified in recent work. The present work was carried using different bioinformatics method in order to determine if Herpesvirus Canine 1 (CHV-1) has a region associated with latency. Our result was the selection of nine sequences candidate of micro RNA (miRNA) (MIREval 2.0 software), and 26 miRNA (miRNAFold v.1.0 software), of them, were selected 14 with real precursors of miRNA, two were found between the RL2 and RS1 genes, one in the RL2 gene and 11 in the RS1 gene. The results showed that the similarities of these regions are very low among the herpesviruses analyzed, so it was not possible to deduce the presence of the LAT gene in canine herpesvirus type 1 with bioinformatics. On the other hand, the comparison showed that the miRNA predicted: chv1-mir-mirnafold-8 has similarity with the ebv-mir-BART7-3p of Epstein-Barr Virus (EBV), in this way, the microRNAs predicted by means of bioinformatic programs met the theoretical requirements of these molecules, however at not having a degree of preservation in other herpesviruses, the expression by CHV-1 in latency cannot be confirmed and it is necessary to identify through experimental tests.展开更多
Herpes simplex virus (HSV) infection in the human body can be latent in neurons for long time and be reactivated leading to recurrence at high rate. Currently there is no effective clinical strategy for the prevention...Herpes simplex virus (HSV) infection in the human body can be latent in neurons for long time and be reactivated leading to recurrence at high rate. Currently there is no effective clinical strategy for the prevention and treatment of the disease relapse. HSV LAT gene is expressed in large quantities and lytic genes are turned off leading to HSV latency. Disruption of the gene expression is thought to cause HSV reactivation and disease relapse. To reveal the essence of HSV latency and reactivation, we summarized and innovatively classified the role, mechanism and transcriptional regulation of LAT in HSV latency and reactivation. This review may have important implications for future studies on HSV latency and reactivation, HSV disease prevention and treatment, and safer and more effective oncolytic HSVs (oHSVs).展开更多
Human mesenchymal stem cells (HMSCs) have emerged as a promising cell type for tissue repopulat-ing and for use as ex vivo gene delivery vehicles. Herpes simplex virus-1 (HSV-1) possesses many vector features, which m...Human mesenchymal stem cells (HMSCs) have emerged as a promising cell type for tissue repopulat-ing and for use as ex vivo gene delivery vehicles. Herpes simplex virus-1 (HSV-1) possesses many vector features, which make it especially suitable for HMSC applications. Here we performed real-time RT-PCR to detect the level of mature microRNA encoded by the HSV-1 latency-associated transcript (microRNA-LAT) in HMSCs cytoplasm with a specific stem loop reverse primer. Three small interfering RNAs (siRNAs) were chemically synthesized towards microRNA-LAT, TGF-β1 and SMAD3. The results demonstrate that HSV-1 and microRNA-LAT prevented HMSCs from undergoing cisplatin-induced apoptosis. In comparison with cells only treated with cisplatin, the apoptosis phenomenon with HSV-1 and microRNA-LAT were markedly reduced. The apoptosis rates of these two groups were both lower (P<0.05) as determined by flow cytometry analysis. The results of RT-PCR and western blotting analysis confirmed that the mRNA and protein levels of TGF-β1 and SMAD3 were significantly decreased with treatment of HSV-1 and microRNA-LAT. Integral TGF-β signalling pathway components were by these means knocked down. Moreover, the levels of the mature microRNA-LAT were decreased in cis-platin-treated HMSCs. In conclusion, HSV-1 and microRNA-LAT exert their anti-apoptotic effect on HMSCs by down-regulation of the TGF-β signalling pathway. Thus HSV-1, having anti-apoptotic effect naturally encoded in its microRNA-LAT, is a good candidate to be developed for HMSC vector.展开更多
Oncolytic virus (OV) is a kind of virus that can preferentially infect and kill tumor cells. The second oncolytic virus drug was oncolytic herpes simplex virus (oHSV) Talimogene Laherparepvec (T-VEC). HSV-1 infectious...Oncolytic virus (OV) is a kind of virus that can preferentially infect and kill tumor cells. The second oncolytic virus drug was oncolytic herpes simplex virus (oHSV) Talimogene Laherparepvec (T-VEC). HSV-1 infectious cell culture protein 34.5 (ICP34.5) and latency-associated transcript (LAT) genes are closely related to virus selective infection and latent infection. Their engineering is essential for constructing efficient and safe oHSV. We summarized the mechanisms of ICP34.5 and LAT in the course of HSV-1 infection and reviewed the engineered oHSVs. We are aimed to provide an insight in developing oHSV in the future.展开更多
Various factors/pathways including hormonal regulation have been suggested to control herpes simplex virus type 1 （HSV-1） latency and reactivation. Our computer analysis identified a DNA repeat containing thyroid ho...Various factors/pathways including hormonal regulation have been suggested to control herpes simplex virus type 1 （HSV-1） latency and reactivation. Our computer analysis identified a DNA repeat containing thyroid hormoneresponsive elements （TRE） in the regulatory region of HSV-1 latency-associated transcript （LAT）. Thyroid hormone （triiodothyronine, T3） functions via its receptor TR （thyroid hormone receptor）, a transcription factor. Present study investigated the roles of TR and T3 in HSV-1 gene expression using cultured neuoroblastoma cell lines. We demonstrated that liganded TR activated LAT transcription, but repressed infected cell protein no. 0 （ICP0） transcription in the presence of LAT TRE. Chromatin immunoprecipitation （CHIP） assays showed that TRs were recruited to LAT TREs independently of T3 and hyperacetylated H4 was associated with the LAT promoter that was transcriptionally active. In addition, ChIP results showed that the chromatin insulator protein CCCTC-binding factor was enriched at the LAT TREs in the presence of TR and T3. In addition, the BRG1 chromatin remodeling complex is found to participate in the T3/TR-mediated LAT activation since overexpression of BRG1 enhanced the LAT transcription and the dominant-negative mutant K785R abolished the activation. This is the first report revealing that TR elicits epigenetic regulation on HSV-1 ICP0 expression in neuronal cells and could have a role in the complex processes of HSV-1 latency/reactivation.展开更多
On entering sensory ganglia,herpes simplex viruses 1(HSV-1)establishes a latent infection with the synthesis of a latency associated transcript(LAT)or initiates productive infection with expression of a set of immedia...On entering sensory ganglia,herpes simplex viruses 1(HSV-1)establishes a latent infection with the synthesis of a latency associated transcript(LAT)or initiates productive infection with expression of a set of immediate early viral proteins.The precise mechanisms how expression of a genes is suppressed during the latency are unknown.One mechanism that has been proposed is illustrated in the case of ICP0,a key immediate early viral regulatory protein.Specifically,the 2 kb LAT intron is complementary to the 30 terminal portion of ICP0 m RNA.To test the hypothesis that accumulation of LAT negatively affects the accumulation of ICP0 m RNA,we inserted a DNA fragment encoding two poly(A)sequences into LAT to early terminate LAT transcript without interrupting the complementary sequence of ICP0 transcript(named as SR1603).Comparisons of the parent(SR1601)and mutant(SR1603)HSV-1 viruses showed the following:Neurons harboring latent SR1603 virus accumulated equivalent amounts of viral DNA but higher amounts of ICP0 m RNA and lower amounts of LAT,when compared to neurons harboring the SR1601 virus.One notable difference between the two viruses is that viral RNA accumulation in explanted ganglia harboring SR1603 virus initiated significantly sooner than that in neurons harboring SR1601 virus,suggesting that ICP0 may act as an activator of viral gene expression in permissive cells.Collectively,these data suggest that increased ICP0 m RNA by suppressed LAT did not affect the establishment of latency in latently infected murine ganglia.展开更多
目的:探讨双秦眼用凝胶对小鼠单疱病毒性角膜炎(HSK)的抗复发作用。方法:用角膜病毒接种法给BALB/c小鼠接种单纯疱疹病毒I型(HSV-1)SM44株,随机分为对照组、无环鸟苷(ACV)组、双秦眼用凝胶组。连续给药2周,停止用药8周后紫外线B(UV-B)...目的:探讨双秦眼用凝胶对小鼠单疱病毒性角膜炎(HSK)的抗复发作用。方法:用角膜病毒接种法给BALB/c小鼠接种单纯疱疹病毒I型(HSV-1)SM44株,随机分为对照组、无环鸟苷(ACV)组、双秦眼用凝胶组。连续给药2周,停止用药8周后紫外线B(UV-B)光照射,诱导HSK复发,观察各组小鼠泪液膜擦拭液的病毒检出率、UV-B光照射前后病变部位体征评分、角膜组织结构、聚合酶链反应(PCR)方法检测三叉神经节(TG)内HSV-1的潜伏,以及反转录聚合酶链反应(RT-PCR)方法检测TG内潜伏相关转录子(LAT)的表达情况。结果:UV-B光照射后,各组检出率均呈不同程度的升高;在照射后7d的泪液膜擦拭液里,除第3-5天双秦眼用凝胶组的泪液膜擦拭液里的病毒检出率高于ACV组外,其余几天,双秦眼用凝胶组的病毒检出率均低于其余两组。但是,比较各组间每天的病毒检出率,均无显著统计学差异;在病变部位体征评分方面,应用双秦眼用凝胶治疗后的小鼠在UV-B光照射诱导后病变程度轻于其余两组(P<0.05);HE染色观察角膜组织结构发现,UV-B光照射后,对照组及ACV组角膜均有不同程度的损伤,双秦眼用凝胶组角膜各层结构大致正常;双秦眼用凝胶组TG内HSV-1的潜伏及LAT的表达均受到抑制;用PCR技术检测各组小鼠TG内HSV-1 DNA的灰度值,结果显示,双秦眼用凝胶优于对照组(P<0.05)及ACV组(P<0.01);应用RT-PCR技术检测并比较各组小鼠TG内LAT m RNA的灰度值,双秦眼用凝胶亦优于对照组(P<0.01)及ACV组(P<0.05)。结论:双秦眼用凝胶在一定程度上可以降低HSK小鼠泪液膜擦拭液的病毒检出率,减轻UV-B光照射后角膜病变体征、保护角膜的组织结构、抑制TG内HSV-1的潜伏和LAT的表达,但双秦眼用凝胶对小鼠HSK的抗复发作用及机制仍需在增加样本量、排除外界因素影响的基础上进一步研究探讨。展开更多
文摘A characteristic common to herpesviruses is the ability to establish a latent infection in the hosts, a transcriptionally active region has detected during latency as well as a set of RNA that are known as Latency Associated Transcripts (LATs), their functions have been clarified in recent work. The present work was carried using different bioinformatics method in order to determine if Herpesvirus Canine 1 (CHV-1) has a region associated with latency. Our result was the selection of nine sequences candidate of micro RNA (miRNA) (MIREval 2.0 software), and 26 miRNA (miRNAFold v.1.0 software), of them, were selected 14 with real precursors of miRNA, two were found between the RL2 and RS1 genes, one in the RL2 gene and 11 in the RS1 gene. The results showed that the similarities of these regions are very low among the herpesviruses analyzed, so it was not possible to deduce the presence of the LAT gene in canine herpesvirus type 1 with bioinformatics. On the other hand, the comparison showed that the miRNA predicted: chv1-mir-mirnafold-8 has similarity with the ebv-mir-BART7-3p of Epstein-Barr Virus (EBV), in this way, the microRNAs predicted by means of bioinformatic programs met the theoretical requirements of these molecules, however at not having a degree of preservation in other herpesviruses, the expression by CHV-1 in latency cannot be confirmed and it is necessary to identify through experimental tests.
文摘Herpes simplex virus (HSV) infection in the human body can be latent in neurons for long time and be reactivated leading to recurrence at high rate. Currently there is no effective clinical strategy for the prevention and treatment of the disease relapse. HSV LAT gene is expressed in large quantities and lytic genes are turned off leading to HSV latency. Disruption of the gene expression is thought to cause HSV reactivation and disease relapse. To reveal the essence of HSV latency and reactivation, we summarized and innovatively classified the role, mechanism and transcriptional regulation of LAT in HSV latency and reactivation. This review may have important implications for future studies on HSV latency and reactivation, HSV disease prevention and treatment, and safer and more effective oncolytic HSVs (oHSVs).
基金the National Natural Science Foundation of China (Grant No. 30772483)Scientific Research Foundation for the Returned Overseas Chinese Scholars, State Education Ministry of P. R. China and the Tianjin Municipal Science and Technology Committee (Grant No. 05YFJMJC03900)
文摘Human mesenchymal stem cells (HMSCs) have emerged as a promising cell type for tissue repopulat-ing and for use as ex vivo gene delivery vehicles. Herpes simplex virus-1 (HSV-1) possesses many vector features, which make it especially suitable for HMSC applications. Here we performed real-time RT-PCR to detect the level of mature microRNA encoded by the HSV-1 latency-associated transcript (microRNA-LAT) in HMSCs cytoplasm with a specific stem loop reverse primer. Three small interfering RNAs (siRNAs) were chemically synthesized towards microRNA-LAT, TGF-β1 and SMAD3. The results demonstrate that HSV-1 and microRNA-LAT prevented HMSCs from undergoing cisplatin-induced apoptosis. In comparison with cells only treated with cisplatin, the apoptosis phenomenon with HSV-1 and microRNA-LAT were markedly reduced. The apoptosis rates of these two groups were both lower (P<0.05) as determined by flow cytometry analysis. The results of RT-PCR and western blotting analysis confirmed that the mRNA and protein levels of TGF-β1 and SMAD3 were significantly decreased with treatment of HSV-1 and microRNA-LAT. Integral TGF-β signalling pathway components were by these means knocked down. Moreover, the levels of the mature microRNA-LAT were decreased in cis-platin-treated HMSCs. In conclusion, HSV-1 and microRNA-LAT exert their anti-apoptotic effect on HMSCs by down-regulation of the TGF-β signalling pathway. Thus HSV-1, having anti-apoptotic effect naturally encoded in its microRNA-LAT, is a good candidate to be developed for HMSC vector.
文摘Oncolytic virus (OV) is a kind of virus that can preferentially infect and kill tumor cells. The second oncolytic virus drug was oncolytic herpes simplex virus (oHSV) Talimogene Laherparepvec (T-VEC). HSV-1 infectious cell culture protein 34.5 (ICP34.5) and latency-associated transcript (LAT) genes are closely related to virus selective infection and latent infection. Their engineering is essential for constructing efficient and safe oHSV. We summarized the mechanisms of ICP34.5 and LAT in the course of HSV-1 infection and reviewed the engineered oHSVs. We are aimed to provide an insight in developing oHSV in the future.
文摘Various factors/pathways including hormonal regulation have been suggested to control herpes simplex virus type 1 （HSV-1） latency and reactivation. Our computer analysis identified a DNA repeat containing thyroid hormoneresponsive elements （TRE） in the regulatory region of HSV-1 latency-associated transcript （LAT）. Thyroid hormone （triiodothyronine, T3） functions via its receptor TR （thyroid hormone receptor）, a transcription factor. Present study investigated the roles of TR and T3 in HSV-1 gene expression using cultured neuoroblastoma cell lines. We demonstrated that liganded TR activated LAT transcription, but repressed infected cell protein no. 0 （ICP0） transcription in the presence of LAT TRE. Chromatin immunoprecipitation （CHIP） assays showed that TRs were recruited to LAT TREs independently of T3 and hyperacetylated H4 was associated with the LAT promoter that was transcriptionally active. In addition, ChIP results showed that the chromatin insulator protein CCCTC-binding factor was enriched at the LAT TREs in the presence of TR and T3. In addition, the BRG1 chromatin remodeling complex is found to participate in the T3/TR-mediated LAT activation since overexpression of BRG1 enhanced the LAT transcription and the dominant-negative mutant K785R abolished the activation. This is the first report revealing that TR elicits epigenetic regulation on HSV-1 ICP0 expression in neuronal cells and could have a role in the complex processes of HSV-1 latency/reactivation.
基金supported by grants from Shenzhen Overseas High-Caliber Peacock Foundation KQTD2015071414385495Shenzhen Science and Innovation Commission Project Grants JCYJ20180306173333907 to Shenzhen International Institute for Biomedical Research。
文摘On entering sensory ganglia,herpes simplex viruses 1(HSV-1)establishes a latent infection with the synthesis of a latency associated transcript(LAT)or initiates productive infection with expression of a set of immediate early viral proteins.The precise mechanisms how expression of a genes is suppressed during the latency are unknown.One mechanism that has been proposed is illustrated in the case of ICP0,a key immediate early viral regulatory protein.Specifically,the 2 kb LAT intron is complementary to the 30 terminal portion of ICP0 m RNA.To test the hypothesis that accumulation of LAT negatively affects the accumulation of ICP0 m RNA,we inserted a DNA fragment encoding two poly(A)sequences into LAT to early terminate LAT transcript without interrupting the complementary sequence of ICP0 transcript(named as SR1603).Comparisons of the parent(SR1601)and mutant(SR1603)HSV-1 viruses showed the following:Neurons harboring latent SR1603 virus accumulated equivalent amounts of viral DNA but higher amounts of ICP0 m RNA and lower amounts of LAT,when compared to neurons harboring the SR1601 virus.One notable difference between the two viruses is that viral RNA accumulation in explanted ganglia harboring SR1603 virus initiated significantly sooner than that in neurons harboring SR1601 virus,suggesting that ICP0 may act as an activator of viral gene expression in permissive cells.Collectively,these data suggest that increased ICP0 m RNA by suppressed LAT did not affect the establishment of latency in latently infected murine ganglia.
文摘目的:探讨双秦眼用凝胶对小鼠单疱病毒性角膜炎(HSK)的抗复发作用。方法:用角膜病毒接种法给BALB/c小鼠接种单纯疱疹病毒I型(HSV-1)SM44株,随机分为对照组、无环鸟苷(ACV)组、双秦眼用凝胶组。连续给药2周,停止用药8周后紫外线B(UV-B)光照射,诱导HSK复发,观察各组小鼠泪液膜擦拭液的病毒检出率、UV-B光照射前后病变部位体征评分、角膜组织结构、聚合酶链反应(PCR)方法检测三叉神经节(TG)内HSV-1的潜伏,以及反转录聚合酶链反应(RT-PCR)方法检测TG内潜伏相关转录子(LAT)的表达情况。结果:UV-B光照射后,各组检出率均呈不同程度的升高;在照射后7d的泪液膜擦拭液里,除第3-5天双秦眼用凝胶组的泪液膜擦拭液里的病毒检出率高于ACV组外,其余几天,双秦眼用凝胶组的病毒检出率均低于其余两组。但是,比较各组间每天的病毒检出率,均无显著统计学差异;在病变部位体征评分方面,应用双秦眼用凝胶治疗后的小鼠在UV-B光照射诱导后病变程度轻于其余两组(P<0.05);HE染色观察角膜组织结构发现,UV-B光照射后,对照组及ACV组角膜均有不同程度的损伤,双秦眼用凝胶组角膜各层结构大致正常;双秦眼用凝胶组TG内HSV-1的潜伏及LAT的表达均受到抑制;用PCR技术检测各组小鼠TG内HSV-1 DNA的灰度值,结果显示,双秦眼用凝胶优于对照组(P<0.05)及ACV组(P<0.01);应用RT-PCR技术检测并比较各组小鼠TG内LAT m RNA的灰度值,双秦眼用凝胶亦优于对照组(P<0.01)及ACV组(P<0.05)。结论:双秦眼用凝胶在一定程度上可以降低HSK小鼠泪液膜擦拭液的病毒检出率,减轻UV-B光照射后角膜病变体征、保护角膜的组织结构、抑制TG内HSV-1的潜伏和LAT的表达,但双秦眼用凝胶对小鼠HSK的抗复发作用及机制仍需在增加样本量、排除外界因素影响的基础上进一步研究探讨。
