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Mitogen-activated Protein Kinase Kinase2, a Novel E2-interacting Protein, Positively Regulates the Replication of Classical Swine Fever Virus

Author: LiaoYaJin
Tutor: ChouHuaJi
School: Chinese Academy of Agricultural Sciences
Course: Preventive Veterinary Medicine
Keywords: Classical swine fever virus (CSFV) E2protein mitogen-activated protein kinase kinase2(MEK2) replication interferon beta
CLC: S852.65
Type: Master's thesis
Year: 2013
Downloads: 2
Quote: 0
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Abstract


Classical swine fever virus (CSFV) is the causative agent of classical swine fever and leads tohuge economic losses to the pig industry. The E2protein is the major envelope glycoprotein of CSFV,which plays a critical role in the CSFV infection of the host cell.To investigate the interactions between the CSFV E2protein and host cellular proteins,E2-interacting proteins were screened from a peripheral blood mononuclear cells (PBMC) cDNA libraryby yeast two-hybrid (Y2H) using E2as a bait. Amomg the17E2-interacting proteins screened,mitogen-activated protein kinase kinase2(MEK2) was chosen for further study. The E2-MEK2interaction was confirmed by GST pulldown, bimolecular fluorescence complementation (BiFC) andlaser confocal assays. The C-terminus of MEK2was identified to be critical for the E2-MEK2interaction.MEK2is an essential kinase in the extracellular signaling-regulated kinase (ERK) pathway. It hasbeen reported that MEK/ERK cascade is involved in the replication of several viruses. Using theanti-pERK antibodies, we demonstrated that CSFV infection activated the MEK/ERK cascade duringthe replication cycle, especially during the early2h post-infection (hpi) and24hpi later.To identify whether the activation is essential to the replication of CSFV, two inhibitors were used,one is U0126, an inhibitor specific for MEK1/2, and the other is PD98059, an MEK1-specific inhibitor.In the supernatants of CSFV-infected PK-15cells treated with U0126, but not with PD98059, CSFVtiters and viral RNA copies were remarkably decreased. The results demonstrated that the activationsignal from MEK2, but not from MEK1, is necessary for the replication of CSFV.Regulation of the cellular cycle and inhibition of the innate immunity are the major mechnismsutilized by viruses to complete the replicaion cycle. To determine the role of MEK2in the interferonbeta (IFN-β) expression, an IFN-β-luciferase reporter system was tested in HEK293T or PK-15cells.The results showed that MEK2s ignificantly inhibited the expression of IFN-β in a dose-dependentmanner.Taken together, the CSFV E2protein interacts with MEK2and activates the MEK/ERK cascade,and inhibition of MEK/ERK cascade significantly decreases the replication of CSFV, possibly viainhibition of the expression of IFN-β by MEK2.

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CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > Animal Medicine ( Veterinary Medicine) > Basic Veterinary Science > Animal Microbiology ( Veterinary Microbiology, ) > Livestock Virology
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