title: Phosphoglycerate kinase inhibits epithelial cell invasion by group B streptococci.
abstract: Group B streptococci (GBS) are opportunistic human pathogens that cause infection and invasive disease in newborns, pregnant women and non-pregnant adults. The internalization of GBS into eukaryotic cells occurs in an actin-microfilament dependent process. The objective of our study was to understand what host cell and/or bacterial factors may be involved in this process. We focused on alpha-actinin, an actin binding protein closely associated with cytoplasmic F-actin in the eukaryotic cell, to determine if it is involved in actin recruitment upon GBS internalization. Initial work revealed that GBS does not recruit alpha-actinin. However, it was found that alpha-actinin antibodies bound to the surface of the GBS, suggesting GBS possess surface-exposed actin binding protein(s). Slide agglutination experiments revealed that when the bacteria were emulsified with F-actin, visible agglutination occurred, further suggesting the presence of an actin binding protein on the GBS cell. Western blot analysis found that anti-alpha-actinin antibodies bound to a 42 kDa protein; mass spectra analysis identified this protein as GBS phosphoglycerate kinase (PGK). Competitive binding assays suggest that the PGK-actin interaction is not a factor in the initial binding of GBS to epithelial cells, however, treating epithelial cells with PGK prior to performing an invasion assay inhibited GBS internalization. This occurred in a dose dependent manner with 10 microg/mL of PGK inhibiting invasion by over 70%, and 50 microg/mL PGK inhibits GBS invasion completely.
authors.affiliation: The Department of Laboratory Medicine and Pathology, The University of Alberta, Edmonton, Alberta, Canada.
authors.forename: Carey-Ann D
authors.initials: CA
authors.lastname: Burnham
authors.forename: Sandra E
authors.initials: SE
authors.lastname: Shokoples
authors.forename: Gregory J
authors.initials: GJ
authors.lastname: Tyrrell
mesh_terms: D000185:Actinin / Q000378:metabolism*; D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000032:analysis / Q000302:isolation & purification; D006367:HeLa Cells; D006801:Humans; D013058:Mass Spectrometry; D008840:Microfilament Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D010735:Phosphoglycerate Kinase / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D013292:Streptococcus agalactiae / Q000472:pathogenicity*
chemical_list: D000199:Actins; D001426:Bacterial Proteins; D008840:Microfilament Proteins; D000185:Actinin; D010735:Phosphoglycerate Kinase