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Invasion of HeLa cells by group B streptococcus requires the phosphoinositide-3-kinase signalling pathway and modulates phosphorylation of host-cell Akt and glycogen synthase kinase-3

¹ The Department of Laboratory Medicine and Pathology, The University of Alberta, Edmonton, AB, Canada
² The National Centre for Streptococcus, The Provincial Laboratory for Public Health (Microbiology), Edmonton, AB, Canada
³ The Department of Medical Microbiology and Immunology, The University of Alberta, Edmonton, AB, Canada

Correspondence
Gregory J. Tyrrell
g.tyrrell{at}provlab.ab.ca

The group B streptococcus (GBS) is an opportunistic bacterial pathogen with the ability to cause invasive disease. While the ability of GBS to invade a number of host-cell types has been clearly demonstrated, the invasion process is not well understood at the molecular level. What has been well established is that modulation of host-cell actin microfilaments is essential for GBS invasion to occur. Phosphoinositide-3 kinase (PI3K) is a key regulator of the cytoskeleton in eukaryotic cells. Our goal in this investigation was to explore the role of the PI3K/Akt signalling pathway in epithelial cell invasion by GBS. The epithelial cell invasion process was mimicked using the HeLa 229 cell-culture model. Treating HeLa cells with chemical inhibitors of PI3K, Akt or Ras prior to bacterial infection inhibited GBS invasion but not attachment; treatment with 30 µM LY294002 (PI3K inhibitor) reduced GBS invasion by 75 %, 20 µM L-6-hydroxymethyl-chiro-inositol 2-(R)-2-O-methyl-3-O-octadecylcarbonate (ICIO) (Akt inhibitor) reduced GBS invasion by 50 %, and 10 µM manumycin A (Ras inhibitor) inhibited GBS invasion by 90 %. Genetic inactivation of the p85 or p110 PI3K subunits in HeLa cells also reduced GBS invasion by 55 and 30 %, respectively. Western blot analysis revealed that phosphorylation of host-cell Akt and glycogen synthase kinase-3 (GSK-3) occurs in response to GBS infection, and that this is mediated upstream by PI3K. Infection of HeLa cells with GBS triggers pro-survival signalling and protects the HeLa cells from camptothecin-induced caspase-3 cleavage. The results from this investigation show that GBS both requires and activates the PI3K/Akt host-cell signalling pathway during invasion of epithelial cells.

A supplementary figure showing the results of experiments to determine whether the inhibitors used affected bacterial membrane permeability to antibiotics is available with the online version of this paper.