Characterization of a new role for plakoglobin in suppressing epithelial cell translocation
Despite advances in the treatment of cancer patients, spread, or metastasis, of tumor cells remains a major cause of death. Although metastasis is a complex process, an important component is cell motility. Acquisition of motility allows tumor cells to breach normal barriers and thereby disseminate. Regulation of motile behavior is poorly understood, but evidence suggests the cadherin family of adhesion molecules plays an important role. The goal of my thesis was to investigate regulation of epithelial cell translocation by cadherins. For my studies, I focused on PAM212 keratinocytes. These cells express high levels of E-cadherin and show suppression of translocation upon contact (STUC). Using repeated light trypsinization and assessment of colony morphology, I selected for variants having a scattered, motile phenotype. As hoped, some variants had reduced expression of E-cadherin (ED cells). Surprisingly, however, some variants retained relatively normal E-cadherin expression but had drastically reduced plakoglobin levels (PD cells). Re-expression of E-cadherin in ED cells, or plakoglobin in PD cells, restored STUC, demonstrating a causal role for each protein. Time-lapse analyses revealed that E-cadherin mediated initial cell-cell contacts, while plakoglobin mediated a subsequent long-term stabilization event. Plakoglobin has known adhesive and signaling functions and could therefore suppress translocation through an adhesive or signaling mechanism. Strong support for a signaling mechanism was provided by: (a) adhesion assays, which revealed no significant difference in adhesiveness of parental PAM212 versus PD cells; (b) time-lapse analyses, which revealed no difference in the ability of parental versus PD cells to form contacts, and; (c) analysis of a deletion-mutant of plakoglobin, which restored STUC despite being unable to participate in desmosome assembly or cadherin adhesion. The most likely candidates for mediating signaling downstream of plakoglobin were TCF/LEF transcription factors, but these were ruled out by: (a) analysis of a dominant-negative TCF/LEF, which failed to render parental cells motile, and; (b) the effects of a deletion-mutant of the plakoglobin-related protein ?-catenin, which restored STUC in PD cells despite being unable to bind these factors. These results reveal a new role for plakoglobin in suppressing epithelial cell translocation, and show that an undescribed signaling pathway underlies this suppression.
School:University of Cincinnati
School Location:USA - Ohio
Source Type:Master's Thesis
Keywords:cell motility plakoglobin cadherin tcf lef signaling movement translocation
Date of Publication:01/01/2001