Dataset Information

Porcine Reproductive and Respiratory Syndrome Virus Infection Induces Stress Granule Formation Depending on Protein Kinase R-like Endoplasmic Reticulum Kinase (PERK) in MARC-145 Cells.

ABSTRACT: Stress granules (SGs) are sites of mRNA storage that are formed in response to various conditions of stress, including viral infections. Porcine reproductive and respiratory syndrome virus (PRRSV) is an Arterivirus that has been devastating the swine industry worldwide since the late 1980s. In this study, we found that infection of PRRSV strain WUH3 (genotype 2 PRRSV) induced stable formation of robust SGs in MARC-145 cells, as demonstrated by the recruitment of marker proteins of SGs, including TIA1, G3BP1, and eIF3?. Treatment with specific inhibitors or siRNAs against the stress kinases that are involved in SG formation revealed that PRRSV induced SG formation through a PERK (protein kinase R-like endoplasmic reticulum kinase)-dependent mechanism. Impairment of SG assembly by concomitant knockdown of the SG marker proteins (TIA1, G3BP1, and TIAR) did not affect PRRSV growth, while significantly enhanced PRRSV-induced NF-?B subunit p65 phosphorylation and inflammatory cytokine production. Taken together, our results demonstrate that PRRSV induces SG formation via a PERK-dependent pathway and that SGs are involved in the signaling pathway of the PRRSV-induced inflammatory response in MARC-145 cells.

SUBMITTER: Zhou Y

PROVIDER: S-EPMC5378712 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Porcine Reproductive and Respiratory Syndrome Virus Infection Induces Stress Granule Formation Depending on Protein Kinase R-like Endoplasmic Reticulum Kinase (PERK) in MARC-145 Cells.

Zhou Yanrong Y Fang Liurong L Wang Dang D Cai Kaimei K Chen Huanchun H Xiao Shaobo S

Frontiers in cellular and infection microbiology 20170404

Stress granules (SGs) are sites of mRNA storage that are formed in response to various conditions of stress, including viral infections. Porcine reproductive and respiratory syndrome virus (PRRSV) is an <i>Arterivirus</i> that has been devastating the swine industry worldwide since the late 1980s. In this study, we found that infection of PRRSV strain WUH3 (genotype 2 PRRSV) induced stable formation of robust SGs in MARC-145 cells, as demonstrated by the recruitment of marker proteins of SGs, in ...[more]

PMID: 28421170

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Project description:MicroRNAs (miRNAs) contribute to gene regulation at the post-transcriptional level and are capable of mRNA silencing by binding to target sites exhibiting high degrees of complementarity. Therefore, cloning host miRNA-recognition sequences into the genome of RNA viruses represents a rational strategy for manipulating viral replication. Here, we performed deep sequencing to obtain small-RNA (sRNA)-expression profiles from in vitro-cultured MARC-145 cells post infection with porcine reproductive and respiratory syndrome virus (PRRSV) and chose six candidate miRNAs of different abundance (miR-21, miR-140-3p, miR-185, miR-26a, miR-505, and miR-199a) for further study. Based on the full-length cDNA clone p7USC, we constructed a number of PRRSV mutants that provided complementary base-pairing target sites for the miRNAs in 3' untranslated regions. Our results showed that all low- and moderate- abundant miRNA-target mutants showed similar growth properties, whereas the highest-abundant miRNA-target mutant blocked both viral transcription and replication. Discontinuous mutations in high-abundant miRNA-target sites subsequently recovered viral viability and propagation. These results demonstrated the copy number of endogenous miRNAs and the extent of sRNA complementarity were key factors to silence potential mRNA expression/translation, thereby determining PRRSV viability. Interestingly, the mutant containing miR-140-target sites (v140-t) showed strong suppression of viral replication from P1 to P3 in vitro, as shown by virus titer, plaque morphology, and qRT-PCR assays. To assess genetic stability, sequencing of v140-t (P1, P3, P5 and P10) revealed spontaneous mutations preferentially located among several nucleotides near the 3' end of the insertion region and corresponding to the "seed region" of miR-140-3p, explaining the induced viral repression and the direction of virus evolution. This approach provided a general silencing strategy for limiting PRRSV replication by endogenous miRNAs in MARC-145 cells.

Dataset Information

Porcine Reproductive and Respiratory Syndrome Virus Infection Induces Stress Granule Formation Depending on Protein Kinase R-like Endoplasmic Reticulum Kinase (PERK) in MARC-145 Cells.

Publications

Porcine Reproductive and Respiratory Syndrome Virus Infection Induces Stress Granule Formation Depending on Protein Kinase R-like Endoplasmic Reticulum Kinase (PERK) in MARC-145 Cells.

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