scholarly journals Accumulation of a 3′-Terminal Genome Fragment in Japanese Encephalitis Virus-Infected Mammalian and Mosquito Cells

2004 ◽  
Vol 78 (10) ◽  
pp. 5133-5138 ◽  
Author(s):  
Kuo-Chih Lin ◽  
Huei-Lan Chang ◽  
Ruey-Yi Chang
Keyword(s):  
Japanese Encephalitis Virus ◽  
Japanese Encephalitis ◽  
Small Rna ◽  
Encephalitis Virus ◽  
Genome Replication ◽  
Rna Molecules ◽  
Genome Fragment ◽  
Mosquito Cells ◽  
Single Positive ◽  
Positive Strand Rna

ABSTRACT Japanese encephalitis virus (JEV) contains a single positive-strand RNA genome nearly 11 kb in length and is not formally thought to generate subgenomic RNA molecules during replication. Here, we report the abundant accumulation of a 3′-terminal 521- to 523-nucleotide (nt) genome fragment, representing a major portion of the 585-nt 3′ untranslated region, in both mammalian (BHK-21) and mosquito (C6/36) cells infected with any of nine strains of JEV. In BHK-21 cells, the viral genome was detected as early as 24 h postinfection, the small RNA was detected as early as 28 h postinfection, and the small RNA was 0.25 to 1.5 times as abundant as the genome on a molar basis between 28 and 48 h postinfection. In C6/36 cells, the genome and small RNA were present 5 days postinfection and the small RNA was 1.25 to 5.14 times as abundant as the genome. The 3′-terminal 523-nt small RNA contains a 5′-proximal stable hairpin (nt 6 to 56) that may play a role in its formation and the conserved flavivirus 3′-cyclization motif (nt 413 to 420) and the 3′-terminal long stable hairpin structure (nt 440 to 523) that have postulated roles in genome replication. Abundant accumulation of the small RNA during viral replication in both mammalian and mosquito cells suggests that it may play a biological role, perhaps as a regulator of RNA synthesis.

scholarly journals 3′ cis-Acting Elements That Contribute to the Competence and Efficiency of Japanese Encephalitis Virus Genome Replication: Functional Importance of Sequence Duplications, Deletions, and Substitutions

2009 ◽  
Vol 83 (16) ◽  
pp. 7909-7930 ◽  
Author(s):  
Sang-Im Yun ◽  
Yu-Jeong Choi ◽  
Byung-Hak Song ◽  
Young-Min Lee
Keyword(s):  
Viral Replication ◽  
Japanese Encephalitis Virus ◽  
Japanese Encephalitis ◽  
Rna Replication ◽  
Encephalitis Virus ◽  
Protein Accumulation ◽  
Genomic Rna ◽  
Human Neuroblastoma ◽  
Lethal Mutant ◽  
Positive Strand Rna

ABSTRACT The positive-strand RNA genome of Japanese encephalitis virus (JEV) terminates in a highly conserved 3′-noncoding region (3′NCR) of six domains (V, X, I, II-1, II-2, and III in the 5′-to-3′ direction). By manipulating the JEV genomic RNA, we have identified important roles for RNA elements present within the 574-nucleotide 3′NCR in viral replication. The two 3′-proximal domains (II-2 and III) were sufficient for RNA replication and virus production, whereas the remaining four (V, X, I, and II-1) were dispensable for RNA replication competence but required for maximal replication efficiency. Surprisingly, a lethal mutant lacking all of the 3′NCR except domain III regained viability through pseudoreversion by duplicating an 83-nucleotide sequence from the 3′-terminal region of the viral open reading frame. Also, two viable mutants displayed severe genetic instability; these two mutants rapidly developed 12 point mutations in domain II-2 in the mutant lacking domains V, X, I, and II-1 and showed the duplication of seven upstream sequences of various sizes at the junction between domains II-1 and II-2 in the mutant lacking domains V, X, and I. In all cases, the introduction of these spontaneous mutations led to an increase in RNA production that paralleled the level of protein accumulation and virus yield. Interestingly, the mutant lacking domains V, X, I, and II-1 was able to replicate in hamster BHK-21 and human neuroblastoma SH-SY5Y cells but not in mosquito C6/36 cells, indicating a cell type-specific restriction of its viral replication. Thus, our findings provide the basis for a detailed map of the 3′ cis-acting elements in JEV genomic RNA, which play an essential role in viral replication. They also provide experimental evidence for the function of 3′ direct repeat sequences and suggest possible mechanisms for the emergence of these sequences in the 3′NCR of JEV and perhaps in other flaviviruses.

scholarly journals Differential miRNA Expression Profiling Reveals Correlation of miR125b-5p with Persistent Infection of Japanese Encephalitis Virus

2021 ◽  
Vol 22 (8) ◽  
pp. 4218
Author(s):  
Chih-Wei Huang ◽  
Kuen-Nan Tsai ◽  
Yi-Shiuan Chen ◽  
Ruey-Yi Chang
Keyword(s):  
Japanese Encephalitis Virus ◽  
Japanese Encephalitis ◽  
Persistent Infection ◽  
Quantitative Polymerase Chain Reaction ◽  
Acute Infection ◽  
Encephalitis Virus ◽  
Luciferase Reporter ◽  
Northern Analysis ◽  
Genome Replication ◽  
Infected Cells

MicroRNAs (miRNAs) play versatile roles in multiple biological processes. However, little is known about miRNA’s involvement in flavivirus persistent infection. Here, we used an miRNA array analysis of Japanese encephalitis virus (JEV)-infected cells to search for persistent infection-associated miRNAs in comparison to acute infection. Among all differentially expressed miRNAs, the miR-125b-5p is the most significantly increased one. The high level of miR-125b-5p in persistently JEV-infected cells was confirmed by Northern analysis and real-time quantitative polymerase chain reaction. As soon as the cells established a persistent infection, a significantly high expression of miR-125b-5p was readily observed. Transfecting excess quantities of a miR-125b-5p mimic into acutely infected cells reduced genome replication and virus titers. Host targets of miR125b-5p were analyzed by target prediction algorithms, and six candidates were confirmed by a dual-luciferase reporter assay. These genes were upregulated in the acutely infected cells and sharply declined in the persistently infected cells. The transfection of the miR125b-5p mimic reduced the expression levels of Stat3, Map2k7, and Triap1. Our studies indicated that miR-125b-5p targets both viral and host sequences, suggesting its role in coordinating viral replication and host antiviral responses. This is the first report to characterize the potential roles of miR-125b-5p in persistent JEV infections.

scholarly journals Early Events in Japanese Encephalitis Virus Infection: Viral Entry

Pathogens ◽  
2018 ◽  
Vol 7 (3) ◽  
pp. 68 ◽  
Author(s):  
Sang-Im Yun ◽  
Young-Min Lee
Keyword(s):  
Cell Surface ◽  
Japanese Encephalitis Virus ◽  
Japanese Encephalitis ◽  
Clinical Manifestations ◽  
Encephalitis Virus ◽  
Host Cells ◽  
Target Cells ◽  
Major Public Health Problem ◽  
Viral Glycoprotein ◽  
Positive Strand Rna

Japanese encephalitis virus (JEV), a mosquito-borne zoonotic flavivirus, is an enveloped positive-strand RNA virus that can cause a spectrum of clinical manifestations, ranging from mild febrile illness to severe neuroinvasive disease. Today, several killed and live vaccines are available in different parts of the globe for use in humans to prevent JEV-induced diseases, yet no antivirals are available to treat JEV-associated diseases. Despite the progress made in vaccine research and development, JEV is still a major public health problem in southern, eastern, and southeastern Asia, as well as northern Oceania, with the potential to become an emerging global pathogen. In viral replication, the entry of JEV into the cell is the first step in a cascade of complex interactions between the virus and target cells that is required for the initiation, dissemination, and maintenance of infection. Because this step determines cell/tissue tropism and pathogenesis, it is a promising target for antiviral therapy. JEV entry is mediated by the viral glycoprotein E, which binds virions to the cell surface (attachment), delivers them to endosomes (endocytosis), and catalyzes the fusion between the viral and endosomal membranes (membrane fusion), followed by the release of the viral genome into the cytoplasm (uncoating). In this multistep process, a collection of host factors are involved. In this review, we summarize the current knowledge on the viral and cellular components involved in JEV entry into host cells, with an emphasis on the initial virus-host cell interactions on the cell surface.

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