RESUMEN
The concept of tRNA recycling has recently emerged from the studies of ribosome-associated quality control. Therein tRNase ZS removes the 2', 3'>p from the ANKZF1-cleaved tRNA and the subsequent TRNT1 action re-generates the intact tRNA. To know the roles of the tRNA recycling in vivo, we investigated how viral infection affects the tRNA recycling system by analyzing the mRNA levels of tRNase ZS and TRNT1. We found that both genes in HeLa cells are upregulated in response to infection of Theiler's mouse encephalitis virus but not to that of an influenza A virus. Upregulation was also observed in cells infected with encephalomyocarditis virus with reduced efficiency. The levels of the IFN-ß mRNA appeared to positively correlate with those of the tRNase ZS and TRNT1 mRNAs. The tRNase ZS gene may be regulated post-transcriptionally in the cells infected with Theiler's mouse encephalitis virus.
Asunto(s)
Endorribonucleasas/genética , Interacciones Huésped-Patógeno/genética , Nucleotidiltransferasas/genética , Procesamiento Postranscripcional del ARN , ARN de Transferencia/genética , Theilovirus/genética , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Virus de la Encefalomiocarditis/genética , Virus de la Encefalomiocarditis/crecimiento & desarrollo , Virus de la Encefalomiocarditis/metabolismo , Endorribonucleasas/metabolismo , Células HeLa , Humanos , Virus de la Influenza A/genética , Virus de la Influenza A/crecimiento & desarrollo , Virus de la Influenza A/metabolismo , Interferón beta/genética , Interferón beta/metabolismo , Isoenzimas/genética , Isoenzimas/metabolismo , Nucleotidiltransferasas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN de Transferencia/metabolismo , Ribosomas/genética , Ribosomas/metabolismo , Theilovirus/crecimiento & desarrollo , Theilovirus/metabolismo , Carga ViralRESUMEN
tRNase ZS (ELAC1) and TRNT1 function in tRNA recycling. Recently, we have shown that these genes are upregulated in the cells infected with Theiler's mouse encephalitis virus (TMEV), implying that tRNA recycling functions in response to viral infection. To address the molecular mechanism underlying the ELAC1 upregulation in the cells infected with TMEV, we performed luciferase assays using various plasmid constructs harboring the ELAC1 promoter region. The luciferase expression from a construct containing the full-length ELAC1 promoter was augmented by TMEV, poly IC, IFN-ß, or IFN-γ. We identified four IFN-stimulated responsible elements (ISREs) in the proximal promoter region. The luciferase expression from the constructs that lack all the ISREs was strongly reduced compared with that from the constructs with the four ISREs in the presence of IFN-ß or IFN-γ. The observation that the ISREs from the ELAC1 promoter are essential for the gene upregulation by IFN-ß or IFN-γ suggests that the ELAC1 gene is upregulated by IFNs.
Asunto(s)
Interferones/farmacología , Regiones Promotoras Genéticas/genética , ARN de Transferencia/genética , Transcripción Genética , Proteínas Supresoras de Tumor/genética , Regulación hacia Arriba/efectos de los fármacos , Antivirales/farmacología , Secuencia de Bases , Células HeLa , Humanos , Interferón beta/farmacología , Interferón gamma/farmacología , ARN de Transferencia/metabolismo , Elementos de Respuesta/genética , Theilovirus/efectos de los fármacos , Theilovirus/fisiología , Regulación hacia Arriba/genéticaRESUMEN
Ribosome-associated quality control (RQC) disassembles aberrantly stalled translation complexes to recycle or degrade the constituent parts. A key step of RQC is the cleavage of P-site tRNA by the endonuclease ANKZF1 (Vms1 in yeast) to release incompletely synthesized polypeptides from ribosomes for degradation. Re-use of the cleaved tRNA for translation requires re-addition of the universal 3'CCA nucleotides removed by ANKZF1. Here, we show that ELAC1 is both necessary and sufficient to remove the 2',3'-cyclic phosphate on ANKZF1-cleaved tRNAs to permit CCA re-addition by TRNT1. ELAC1 activity is optimized for tRNA recycling, whereas ELAC2, the essential RNase Z isoform in eukaryotes, is required to remove 3' trailers during tRNA biogenesis. Cells lacking ELAC1 specifically accumulate unrepaired tRNA intermediates upon the induction of ribosome stalling. Thus, optimal recycling of ANKZF1-cleaved tRNAs in vertebrates is achieved through the duplication and specialization of a conserved tRNA biosynthesis enzyme.