Influenza A Virus-Induced Early Activation of ERK and PI3K Mediates V-ATPase-dependent Intracellular pH Change Required for Fusion

Cell Microbiol. 2011 Apr;13(4):587-601. doi: 10.1111/j.1462-5822.2010.01556.x. Epub 2010 Dec 29.

Abstract

The vacuolar (H+)-ATPases (V-ATPases) facilitate the release of influenza A virus (IAV) genome into the cytoplasm by acidifying the endosomal interior. The regulation of V-ATPases by signalling pathways has been demonstrated in various model systems. However, little is known about signalling-regulated V-ATPase activation during IAV infection. Here we show that V-ATPase activity is elevated during infection of cell monolayers with IAV, as measured by intracellular pH change, via a mechanism mediated by extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3-kinase (PI3K). Inhibition of IAV-induced early activation of these kinases reduced V-ATPase activity and the acidification of intracellular compartments in infected cells. IAV-activated ERK and PI3K appear to interact directly, and they colocalize with the E subunit of V-ATPase V1 domain. Further, siRNAs targeting the E2 subunit isoform significantly reduced virus titres. Interestingly, suppression of PI3K early activation, but not that of ERK or V-ATPase, negatively affected virus internalization, suggesting the involvement of the pathway in earlier, V-ATPase-independent infection-promoting events. Cell treatment with a V-ATPase-specific inhibitor impaired the nuclear localization of incoming viral ribonucleoproteins, inhibiting replication/transcription of viral RNAs. These findings highlight the importance of IAV-induced ERK and PI3K early activation as signalling mediators in V-ATPase-stimulated endosomal acidification required for fusion.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Enzyme Activation
  • Extracellular Signal-Regulated MAP Kinases / metabolism*
  • Humans
  • Hydrogen-Ion Concentration
  • Influenza A virus / enzymology*
  • Influenza A virus / genetics
  • Influenza A virus / physiology*
  • MAP Kinase Kinase Kinases / metabolism
  • Phosphatidylinositol 3-Kinase / metabolism*
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • Protein Subunits / genetics
  • Protein Subunits / metabolism
  • Proto-Oncogene Proteins c-akt / metabolism
  • RNA, Small Interfering / metabolism
  • Signal Transduction / physiology
  • Vacuolar Proton-Translocating ATPases / genetics
  • Vacuolar Proton-Translocating ATPases / metabolism*
  • Virus Internalization*
  • raf Kinases / metabolism

Substances

  • Protein Isoforms
  • Protein Subunits
  • RNA, Small Interfering
  • Phosphatidylinositol 3-Kinase
  • Proto-Oncogene Proteins c-akt
  • raf Kinases
  • Extracellular Signal-Regulated MAP Kinases
  • MAP Kinase Kinase Kinases
  • Vacuolar Proton-Translocating ATPases