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Title: The role of tetraspanins in multinucleated giant cell formation induced by Burkholderia thailandensis
Author: Elgawidi, Atiga
ISNI:       0000 0004 5990 141X
Awarding Body: University of Sheffield
Current Institution: University of Sheffield
Date of Award: 2016
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The tetraspanins are a superfamily of transmembrane proteins with a wide distribution in multicellular organisms. Tetraspanins have been involved in many cellular functions including adhesion, migration and the immune response. Tetraspanins are considered as cell membrane organizers and their function is likely to be due the ability to interact with cell membrane proteins. These interactions lead to them forming a large extended network known as tetraspanin enriched microdomains (TEM) or the tetraspanin web. There is growing evidence for the role of some tetraspanins in various fusion events including sperm-egg fusion, viral-cell fusion and monocyte fusion. Monocyte/macrophage fusion is associated with chronic inflammation and is thought to be important for immune defence. Monocyte fusion leading to multinucleated giant cell formation (MNGC) has been linked with the pathogenesis of Burkholderia pseudomallei a causative agent of melioidosis disease, a severe invasive disease endemic in south Asia and North Australia. However the mechanism of B.p-induced MNGC formation is unclear. The role of tetraspanins in MNGC formation has been demonstrated previously. Antibodies to CD9 and CD81 enhanced the fusion of con A stimulated peripheral blood monocytes and the deletion of CD9/CD81 resulted in enhanced MNGC formation in vivo and in vitro. In addition recombinant proteins of tetraspanin large extracellular domain (EC2) of CD9 inhibited MNGC formation, suggesting that CD9 and CD81 are negatively regulating monocyte fusion (Takeda et al., 2003). These finding were confirmed by our research group, who also reported that CD63 is involved in this process as anti CD63 antibodies and CD63 EC2 proteins inhibited Con A-stimulated monocyte fusion whereas CD151 EC2 and CD82 EC2 had no effect (Parthasarathy et al., 2009). This thesis describes attempts made to investigate the role of tetraspanins in MNGC formation induced by B.thailandensis, a non-pathogenic species that is closely related to B.pseudomallei and widely used as a model to study B.p features. Using antibodies to tetraspanins, recombinant EC2 proteins, and tetraspanin deficient cell lines we found that tetraspanins are involved in B.thailandensis-induced cell fusion. Our data is partly in line with previous findings: antibodies to CD9 and CD81 enhanced MNGC formation, whereas CD9 EC2 and CD63 EC2 proteins inhibit fusion. In contrast with other results mentioned above, we found that CD81 EC2 proteins also inhibited B.t-induced fusion. The deletion of CD9 and CD82 resulted in an enhanced MNGC formation, confirming that CD9 negatively promotes B.t-induced cell fusion and suggesting the involvement of CD82 in this process. Further investigation revealed that the absence of tetraspanins could affect the expression level of other cell surface molecules that have been implicated in fusion at the protein level but not at the gene level, which may be relevant to their role as cell membrane organisers. The role of some of these molecules in B.t-induced fusion was also investigated in this study. Several studies showed that Burkholderia induced MNGC formation is facilitated by bacterial effectors, which bring the plasma membrane of infected cells to very close position where they can spread from one cell to another. It is likely that the bacteria regulate this process by affecting the expression of host cell membrane molecules including tetraspanins.
Supervisor: Partridge, Lynda Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available