RESUMO
Herpes zoster (HZ) remains a significant health burden with millions of cases in North America and Europe annually. HZ is frequently followed by long-term pain or post-herpetic neuralgia (PHN). Although effective vaccines for HZ are available, currently used nucleotide analogues often have limited effectiveness against HZ and especially PHN, so there remains a need for additional antiviral therapies for HZ. We recently identified a population of small non-coding RNA (sncRNA) encoded by Varicella Zoster Virus (VZV) and showed that single locked-nucleic acid antagonists (LNAA) to some sncRNA can modulate VZV replication in cell culture. In this work, we explored the antiviral effects of combinations of LNAA oligonucleotides targeting VZVsncRNA. Combinations of LNAA targeting three VZVsncRNA encoded in and near a critical viral regulatory gene were additive, achieving 96 % reduction in virus growth in a cell line. VZV growth was also inhibited by more than 90 % in primary human skin fibroblast cultures by individual and combinations of LNAA to VZVsncRNA. The inhibition by VZVsncRNA was specific and not a consequence of innate immune responses since LNAA to a different VZVsncRNA enhanced VZV growth. Targeted VZVsncRNA lack homologous sequences in the human transcriptome suggesting that LNAA to them would have reduced cytotoxicity if used as therapeutics. These results support further development of oligonucleotides targeting VZVsncRNA as a novel treatment for HZ.
Assuntos
Antivirais/farmacologia , Herpesvirus Humano 3/efeitos dos fármacos , Ácidos Nucleicos/farmacologia , Pequeno RNA não Traduzido/genética , Fibroblastos , Herpes Zoster/tratamento farmacológico , Herpesvirus Humano 3/fisiologia , Humanos , Oligonucleotídeos/síntese química , Pequeno RNA não Traduzido/antagonistas & inibidores , Replicação Viral/efeitos dos fármacosRESUMO
Most herpesviruses use both host and viral small non-coding RNAs (sncRNA), especially microRNA, to modulate infection. Bioinformatic analyses of NGS data obtained from Varicella Zoster virus (VZV)-infected cells predicted 24 VZVsncRNA, seven of which were confirmed to be expressed in infected fibroblasts and neurons using stem-loop quantitative reverse transcription PCR (SL-PCR). We here assayed for the expression of all 24 of the bioinformatically predicted VZVsncRNA in cells productively infected by VZV using SL-PCR. 23 of the 24 predicted sequences were detected in VZV-infected ARPE19 cells and 19 of the 24 sequences in infected human neurons generated by two methods from embryonic stem cells. We also show that blocking one of two newly-tested VZV-encoded sncRNA using locked nucleotide antagonists significantly increased viral replication. These findings suggest that further study of VZV encoded sncRNA could elucidate an additional level of regulation of the life cycle of this pathogenic human herpesvirus.
Assuntos
Células Epiteliais/virologia , Herpesvirus Humano 3/genética , Neurônios/virologia , Pequeno RNA não Traduzido/genética , RNA Viral/genética , Células Cultivadas , Biologia Computacional , Perfilação da Expressão Gênica , Herpesvirus Humano 3/fisiologia , Humanos , Pequeno RNA não Traduzido/antagonistas & inibidores , RNA Viral/antagonistas & inibidores , Replicação Viral/genéticaRESUMO
Transfer-RNA-derived small RNAs (tsRNAs; also called tRNA-derived fragments) are an abundant class of small non-coding RNAs whose biological roles are not well understood. Here we show that inhibition of a specific tsRNA, LeuCAG3'tsRNA, induces apoptosis in rapidly dividing cells in vitro and in a patient-derived orthotopic hepatocellular carcinoma model in mice. This tsRNA binds at least two ribosomal protein mRNAs (RPS28 and RPS15) to enhance their translation. A decrease in translation of RPS28 mRNA blocks pre-18S ribosomal RNA processing, resulting in a reduction in the number of 40S ribosomal subunits. These data establish a post-transcriptional mechanism that can fine-tune gene expression during different physiological states and provide a potential new target for treating cancer.
Assuntos
Pequeno RNA não Traduzido/genética , RNA de Transferência de Leucina/genética , Proteínas Ribossômicas/biossíntese , Ribossomos/genética , Ribossomos/metabolismo , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Pareamento de Bases , Sequência de Bases , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/terapia , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Feminino , Células HCT116 , Células HEK293 , Células HeLa , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/terapia , Masculino , Camundongos , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/farmacologia , Oligonucleotídeos Antissenso/uso terapêutico , Biossíntese de Proteínas/efeitos dos fármacos , Biossíntese de Proteínas/genética , RNA Mensageiro/química , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Ribossômico 18S/genética , RNA Ribossômico 18S/metabolismo , Pequeno RNA não Traduzido/antagonistas & inibidores , RNA de Transferência de Leucina/antagonistas & inibidores , Proteínas Ribossômicas/genética , Subunidades Ribossômicas Menores de Eucariotos/metabolismo , Ribossomos/efeitos dos fármacos , Especificidade por Substrato/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Viral infection initiates an array of changes in host gene expression. Many viruses dampen host protein expression and attempt to evade the host anti-viral defense machinery. Host gene expression is suppressed at several stages of host messenger RNA (mRNA) formation including selective degradation of translationally competent messenger RNAs. Besides mRNAs, host cells also express a variety of noncoding RNAs, including small RNAs, that may also be subject to inhibition upon viral infection. In this review we focused on different ways viruses antagonize coding and noncoding RNAs in the host cell to its advantage.
