Giant cell formation by macrophages and lung epithelial cells: A unique method of cell-cell infection used by B. thailandensis involves tetraspanins

Burkholderia thailandensis is widely used as a non-pathogenic model of Burkholderia pseudomallei, the causative agent of melioidosis, a disease with a 90% mortality rate if untreated. One of the histopathological features of melioidosis is the presence of multinucleate giant cells (MGC). MGC formation occurs when the plasma membrane of infected cells become closely positioned with other cells, which then become attached and fuse together. Many studies have described how effector systems of B. thailandensis can induce the formation of MGC. It seems that these bacterial effectors can regulate this process by affecting the expression of mammalian membrane proteins, including the tetraspanins (Tspans) superfamily. Tspans are a large family of membrane proteins that bind partner proteins to form Tspan-enriched microdomains (TEM) that have many biological roles, including cell fusion, adhesion, and bacterial infection. Mammalian cells express 33 Tspans, but their specific functions during MGC formation and pathogen infection are not fully understood. Here, we have attempted to define the roles of all Tspans and some Tspan-partner proteins in B. thailandensis infection and/or MGC formation. We found that 5 Tspans are specifically involved in MGC formation induced by B. thailandensis: Tspan-2, Tspan-5, Tspan-13, CD81, and CD9. 3 Tspan-partner proteins are also specifically involved: ADAM10, CD98, and CD172α. Using antibodies, an inhibitory peptide derived from CD9 and knockouts and knockdowns of Tspans and their partners, we attempted to elucidate the roles of these molecules in MGC formation. It was observed that CD9, CD81, CD172α, and ADAM10 have negative roles in MGC formation induced by B. thailandensis whereas Tspan-2 and Tspan-13 play positive roles and could also be required for B. thailandensis infection. CD98 also has a positive role in MGC formation but has no role in B. thailandensis infection. It was also found that the peptide derived from CD9 could reduce the total number of bacteria of B. thailandensis, and internalisation after 2 and 18hr.