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Protein kinase A-dependent step(s) in hepatitis C virus entry and infectivity

Farquhar, Michelle J and Harris, Helen J and Diskar, Mandy and Jones, Sarah and Mee, Christopher J and Nielsen, Søren U and Brimacombe, Claire L and Molina, Sonia and Toms, Geoffrey L and Maurel, Patrick and Howl, John and Herberg, Friedrich W and van IJzendoorn, Sven C D and Balfe, Peter and McKeating, Jane A (2008) Protein kinase A-dependent step(s) in hepatitis C virus entry and infectivity. Journal of virology, 82 (17). pp. 8797-811. ISSN 1098-5514

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URL of Published Version: http://jvi.asm.org/content/82/17/8797.abstract?sid=fc8d9fc0-1590-4348-8d8a-736df906a7c2

Identification Number/DOI: 10.1128/JVI.00592-08

Viruses exploit signaling pathways to their advantage during multiple stages of their life cycle. We demonstrate a role for protein kinase A (PKA) in the hepatitis C virus (HCV) life cycle. The inhibition of PKA with H89, cyclic AMP (cAMP) antagonists, or the protein kinase inhibitor peptide reduced HCV entry into Huh-7.5 hepatoma cells. Bioluminescence resonance energy transfer methodology allowed us to investigate the PKA isoform specificity of the cAMP antagonists in Huh-7.5 cells, suggesting a role for PKA type II in HCV internalization. Since viral entry is dependent on the host cell expression of CD81, scavenger receptor BI, and claudin-1 (CLDN1), we studied the role of PKA in regulating viral receptor localization by confocal imaging and fluorescence resonance energy transfer (FRET) analysis. Inhibiting PKA activity in Huh-7.5 cells induced a reorganization of CLDN1 from the plasma membrane to an intracellular vesicular location(s) and disrupted FRET between CLDN1 and CD81, demonstrating the importance of CLDN1 expression at the plasma membrane for viral receptor activity. Inhibiting PKA activity in Huh-7.5 cells reduced the infectivity of extracellular virus without modulating the level of cell-free HCV RNA, suggesting that particle secretion was not affected but that specific infectivity was reduced. Viral particles released from H89-treated cells displayed the same range of buoyant densities as did those from control cells, suggesting that viral protein association with lipoproteins is not regulated by PKA. HCV infection of Huh-7.5 cells increased cAMP levels and phosphorylated PKA substrates, supporting a model where infection activates PKA in a cAMP-dependent manner to promote virus release and transmission.

Type of Work:Article
Date:2008 (Publication)
School/Faculty:Schools (1998 to 2008) > School of Medicine
Department:Immunity and Infection
Subjects:R Medicine (General)
QR355 Virology
QR180 Immunology
QR Microbiology
Institution:University of Birmingham
Copyright Holders:American Society for Microbiology
ID Code:474
Refereed:YES
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