Fig 4.

Perinuclear distribution of internalized A. actinomycetemcomitans OMV proteins in HeLa cells and HGF. A. actinomycetemcomitans strain D7SS (WT) and D7SS cdtABC (cdtABC) OMVs and PBS (buffer) were used to treat HeLa cells (A) and HGF (B) for 72 h. After treatment, cells were fixed and incubated with an FITC-conjugated antibody specific for E. coli OmpA or for whole cells of A. actinomycetemcomitans serotype a, as indicated. Actin filaments and nuclei were then stained with phalloidin (green) and DAPI (blue), respectively. Bound antibodies were detected with TRITC (red). The left panels show merged images from staining with the three dyes. The right panels show images from TRITC staining only. Confocal Z-stack projections are included in all images. The cross hairs indicate the positions of the xz and yz planes. Magnification, ×1,000. Bars = 10 μm. (C) Assessment of the cytoplasmic localization of OmpA. HeLa cells were treated with strain D7SS or D7SS cdtABC OMVs for 72 h. Shown are the means ± standard deviations (SD) for the cytoplasmic/nuclear ratio of red fluorescence from 20 cells from three experiments.