UBC Theses and Dissertations
The role of Connexin43 carboxyl terminal domain in regulating various signaling-induced cytoskeleton-dependent cellular processes in B-lymphocytes Pournia, Farnaz
Cellular processes requiring cytoskeleton remodeling such as cell adhesion, cell spreading, protrusion extension, immune synapse formation, cell motility and directional migration are essential for normal B lymphocyte (B cell) development and for immune responses. These cellular processes occur downstream of different signaling pathways that are tightly regulated and dysregulation of the signaling pathways are associated with pathological conditions such as immune deficiencies, autoimmune diseases, and B cell malignancies. Previous work by our group showed the importance of the carboxyl tail (CT) domain of the most ubiquitously expressed member of the gap junction family, connexin43 (Cx43) in supporting BCR-mediated membrane spreading and LFA-1 mediated adhesion of B cells. However, whether the CT domain can support various cytoskeleton-dependent processes independent of the other domains of the protein was not known. Additionally, previous studies in the Matsuuchi lab revealed the importance of the tyrosine residues of the Cx43 CT as potential phosphorylation sites for supporting BCR-mediated spreading. The importance of other potential phosphorylation sites of the Cx43 CT such as the serine residues was not previously explored. In this dissertation, using chimeric proteins containing the CT domain of Cx43 we were able to express the CT domain of Cx43 at the plasma membrane, which was first of its kind. Our results showed that the CT domain of Cx43, by itself supports BCR-mediated spreading. Next, we showed site-specific phosphorylation of the Cx43 CT at Y247, S255, S279, S282 and S368 residues. Using a mutational approach, we showed the importance of S279 and S282 residues, located within a SH3-biniding domain, for BCR-mediated spreading. Lastly, we showed that Cx43 differentially affects the activation of various signaling molecules and the downstream cytoskeleton-dependent cellular processes. These findings suggest that the CT of Cx43 is modified and differentially affects various signaling induced cytoskeleton-dependent processes in B cells. These results provide a novel understanding of the functions of Cx43 CT for B cell responses that are important for immune functions. These findings could be important for development of novel therapeutic approaches for B cell related disease and could possibly be extended to other cell types.
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