Plasma membrane protrusions mediate host cell-cell fusion induced by Burkholderia thailandensis

Cell-cell fusion is important for biological processes including fertilization, development, immunity, and microbial pathogenesis. Bacteria in the pseudomallei group of Burkholderia species, including B. thailandensis, spread between host cells by inducing cell-cell fusion. Previous work showed that B. thailandensis-induced cell-cell fusion requires intracellular bacterial motility and a bacterial protein secretion apparatus called the type VI secretion system-5 (T6SS-5), including the T6SS-5 protein VgrG5. However, the cellular level mechanism and T6SS-5 proteins important for bacteria-induced cell-cell fusion remained incompletely described. Using live cell imaging, we found bacteria used actin-based motility to push on the host cell plasma membrane to form plasma membrane protrusions that extended into neighboring cells. Then, membrane fusion occurred within these membrane protrusions, either proximal to the bacterium at the tip or elsewhere within a protrusion. Expression of VgrG5 by bacteria within membrane protrusions was required to promote cell-cell fusion. Furthermore, a second predicted T6SS-5 protein, TagD5, was also required for cell-cell fusion. In the absence of VgrG5 or TagD5, bacteria in plasma membrane protrusions were engulfed into neighboring cells. Our results suggest that the T6SS-5 effectors VgrG5 and TagD5 are secreted within membrane protrusions and act locally to promote membrane fusion.