Cholesterol and host cell surface proteins contribute to cell-cell fusion induced by the Burkholderia type VI secretion system 5

Abstract

Following escape into the cytoplasm of host cells, Burkholderia pseudomallei and the related species Burkholderia thailandensis employ the type VI secretion system 5 (T6SS-5) to induce plasma membrane fusion with an adjacent host cell. This process leads to the formation of multinucleated giant cells and facilitates bacterial access to an uninfected host cell in a direct manner. Despite its importance in virulence, the mechanism of the T6SS-5 and the role of host cell factors in cell-cell fusion remain elusive. To date, the T6SS-5 is the only system of bacterial origin known to induce host-cell fusion. To gain insight into the nature of T6SS-5-stimulated membrane fusion, we investigated the contribution of cholesterol and proteins exposed on the host cell surface, which were shown to be critically involved in virus-mediated giant cell formation. In particular, we analyzed the effect of host cell surface protein and cholesterol depletion on the formation of multinucleated giant cells induced by B. thailandensis. Acute protease treatment of RAW264.7 macrophages during infection with B. thailandensis followed by agarose overlay assays revealed a strong reduction in the number of cell-cell fusions compared with EDTA treated cells. Similarly, proteolytic treatment of specifically infected donor cells or uninfected recipient cells significantly decreased multinucleated giant cell formation. Furthermore, modulating host cell cholesterol content by acute cholesterol depletion from cellular membranes by methyl- β-cyclodextrin treatment or exogenous addition of cholesterol impaired the ability of B. thailandensis to induce cell-cell fusions. The requirement of physiological cholesterol levels suggests that the membrane organization or mechanical properties of the lipid bilayer influence the fusion process. Altogether, our data suggest that membrane fusion induced by B. pseudomallei and B. thailandensis involves a complex interplay between the T6SS-5 and the host cell.

Involvement of lipid microdomains in human endothelial cells infected by Streptococcus agalactiae type III belonging to the hypervirulent ST-17

“…(8) Cholesterol-enriched membrane microdomains may provide a platform to concentrate receptors on the host cell membrane. (17) Our data support the notion that lipid rafts on the plasma membrane of HUVEC cells facilitate entry of S. agalactiae GBS90356 strain. The significant decrease in cytoadhesion of GBS90356 strain to human endothelial cells treated with MβCD indicated that cholesterol depletion from the cell membrane perturbed the attachment of bacteria and altered the GBS90356 entry at post-binding steps.…”

Multinucleated Giant Cell Formation as a Portal to Chronic Bacterial Infections

“…An intact T6SS-5 is necessary for cell fusion but the identity and function of other delivered effector molecules, beyond VgrG-5, remains to be discovered. On the host side, membrane cholesterol and protein content appear to be important for proper membrane fusion, and this is more than likely associated with optimal membrane thickness for T6SS utilization [ 30 ]. In another study, MNGC formation was blocked by antibodies that targeted host surface molecules [ 31 ].…”

Multinucleated Giant Cell Formation as a Portal to Chronic Bacterial Infections

“…A completely intact T6SS-5 is necessary for cell fusion but the identity and function of any delivered effector molecules, beyond VgrG-5, remains to be discovered. On the host side, membrane cholesterol and protein content appear to be important for proper membrane fusion and this is more than likely associated with optimal membrane thickness for T6SS firing and membrane penetration [28]. Another study was able to block MNGC formation by using antibodies that targeted host surface molecules [29].…”