Altered cytoskeletal regulation impacts numerous physiological phenomena: cell motility, apoptosis, oncogenic transformation and parasitic infection. The protein tyrosine phosphatase (PTP)-PEST contains multiple motifs mediating its recruitment to signalling components, and is required for actin filament organization. However, little is known regarding either the importance of PTP-PEST subcellular localization, or the role of PTP-PEST in either parasitic infection or apoptosis. My doctoral research was therefore focussed on elucidating the effect of subcellular distribution on PTP-PEST activity, specifically with respect to regulation of p130Cas (a PTP-PEST substrate), as well as on the involvement of PTP-PEST in both host-pathogen relations and apoptosis. First, PTP-PEST was found both within the cytosol and at the plasma membrane. Using PTP-PEST -/- rescued cell lines, I observed that tyrosine phosphorylation-dependent p130Cas interactions were controlled primarily by cytosolic PTP-PEST. Secondly, infection of fibroblasts with Leishmania major was observed to induce dramatic actin rearrangements, and to alter the phosphorylation state of numerous proteins. Importantly, both PTP-PEST and p130Cas were processed by the parasitic protease GP63 during infection. GP63 was also required for the cleavage of additional host proteins: cortactin, TC-PTP and caspase-3. Of note, Leishmania parasites mediated p38 inactivation, correlating with the proteolysis of its upstream activator TAB1, in a GP63dependent manner. These results indicate that GP63 plays a key role in a number of biochemical events, potentially contributing to Leishmania infectivity. Finally, PTP-PEST was found to relocalize to the edges of retracting membrane ruffles of apoptotic cells. Surprisingly, PTP-PEST was specifically cleaved by caspase-3 at the 549DSPD motif during apoptosis; leading to modification of catalytic activity and scaffolding properties, and sensitizing cells to Fas-mediated detachment. As this data demonstrated a potential role for caspase cleavage in PTP regulation, I also investigated the presence of conserved putative caspase-c1eavage sites in other family members. In summary, the data presented herein links PTP-PEST with various biological processes: oncogenic signalling, host-pathogen interactions, and apoptosis. In addition to demonstrating the involvement of PTP-PEST in diverse signalling pathways, these studies underscore the importance of subcellular localization and proteolysis in the regulation of this PTP.

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