Early sequence of events triggered by the interaction of Neisseria meningitidis with endothelial cells
Version of Record online: 27 DEC 2013
© 2013 John Wiley & Sons Ltd
Volume 16, Issue 6, pages 878–895, June 2014
How to Cite
Soyer, M., Charles-Orszag, A., Lagache, T., Machata, S., Imhaus, A.-F., Dumont, A., Millien, C., Olivo-Marin, J.-C. and Duménil, G. (2014), Early sequence of events triggered by the interaction of Neisseria meningitidis with endothelial cells. Cellular Microbiology, 16: 878–895. doi: 10.1111/cmi.12248
- Issue online: 16 MAY 2014
- Version of Record online: 27 DEC 2013
- Accepted manuscript online: 10 DEC 2013 06:04AM EST
- Manuscript Accepted: 3 DEC 2013
- Manuscript Revised: 12 NOV 2013
- Manuscript Received: 19 JUN 2013
- Fondation pour la Recherche Médicale FRM
- French ministry for research and higher education
- Avenir INSERM Starting Grant
- Région Ile de France
- European Research Council
Fig. S1. Actin and membrane accumulation in ΔpilV and ΔpilT strains. Endothelial cells transfected with actin-mCherry or Farnesyl-GFP were infected with aggregates formed by the pilV- and pilT-deficient strains (ΔpilV, ΔpilT).
A. Kinetics of actin accumulation in the ΔpilV (green curve) and ΔpilT (red curve) strains.
B. Kinetics of membrane marker accumulation in the ΔpilV (green curve) and ΔpilT (red curve) strains.
Fig. S2. Complementation of the ΔpilV and ΔpilT mutations. Strains expressing wild-type copies of pilV and pilT under the control of the lac promoter in the presence of 1 mM and 0.05 mM of IPTG respectively were used to infect endothelial cells transfected with ezrin-GFP.
A. Kinetics of ezrin accumulation in the pilVind (red curve).
B. Kinetics of ezrin accumulation in the pilTind (red curve).
|cmi12248-sup-0004-si.pdf||2440K||Fig. S3. Evaluation of the effect of ERM knock-down by immunofluorescence. Expression of ezrin and of CD44 as a control were visualized by immunofluorescence in cells treated with ERM siRNA (SiERM) and cells treated with control siRNA. The scale bar corresponds to 20 μm.|
|cmi12248-sup-0005-si.mp4||361K||Movie S1. Meningococcus growth as a microcolony is accompanied by ezrin accumulation. Ezrin-GFP transfected cells were infected by individual mCherry-expressing meningococci. Following initial adhesion (white arrows), behaviour of ezrin underneath the nascent microcolony was monitored over a 4 h period. As the microcolony reached a size of about 10 diplococci (30 min), ezrin started to accumulate and the cortical plaque size increased with the size of the microcolony. The scale bar corresponds to 20 μm.|
|cmi12248-sup-0006-si.mp4||527K||Movie S2. Ezrin accumulation underneath pre-formed meningococcus aggregates. Ezrin-GFP transfected cells (middle panel) were infected with pre-formed mCherry-expressing meningococcus aggregates (left panel). Ezrin accumulation was monitored over 10 min and started immediately after contact of the aggregate with the cell. Right panel shows the merged images. Scale bar corresponds to 20 μm.|
|cmi12248-sup-0007-si.mp4||1650K||Movie S3. Delayed plasma membrane remodelling by the ΔpilT strain. Endothelial cells were transfected with ezrin-GFP and infected with the ΔpilT strain for a period of 4 h. The arrow points to a bacterial aggregate that triggers a detectable response only after 1 h and 15 min. Bacteria (left), ezrin-GFP (middle) and merge (right) are shown on the movie. Scale bar corresponds to 20 μm.|
|cmi12248-sup-0008-si.mp4||601K||Movie S4. Treatment of endothelial cells with cytochalasin D leads to disruption of actin cytoskeleton. Endothelial cells were co-transfected with LifeAct-mCherry, allowing direct visualization of the actin cytoskeleton, and with ezrin-GFP. Actin cytoskeleton behaviour following cytochalasin D treatment was monitored over 20 min. Soon after drug addition, actin fibres started to depolymerize and led to a punctuate labelling of actin throughout the cell. Scale bar corresponds to 20 μm.|
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