Background: Pancreatic cancer is a highly invasive malignancy. Ezrin, a plasma membrane-cytoskeletal linker protein, is associated with the invasive behaviour of cancers.
Aims: Elucidate a possible molecular mechanism for the invasive phenotype.
Methods: Using a combination of techniques, such as Western blotting, co-immuno-precipitation, confocal and light microscopy, invasion & adhesion assays, organotypic cultures and human samples as well as RNAi and expression of various mutant Ezrin constructs, we have dissected out the dynamic molecular nature of podosomes in pancreatic cancer.
Results: Podosome and podosomal rosette formation in pancreatic carcinoma (PaCa3) cells is Ezrin dependent and associated with adhesion to Fibronectin with subsequent digestion of this substrate. Ezrin binds to increasing amounts of Cortactin during formation of the podosomal rosette, with the C-terminal region, specifically the actin binding domain, mediating this molecular linkage. Further, we show that phosphorylation of Tyr353 and Thr567 sites on Ezrin (conventionally shown to translocate Ezrin to plasma membrane) is not required for podosome formation. The podosomal rosette is revealed to be a highly dynamic and transient structure, which can metamorphose into other cellular processes, such as filopodia or lamellipodia, and thereby enable epithelial cancer cells to 'palpate' the underlying substrate and modify their cytoskeletal behaviour accordingly. In human tumour tissues and organotypic cultures, specific sub-cellular expression of Ezrin (basal membranous; cellular processes invading stroma) in pancreatic cancer cells can be correlated with tumour progression and disease free survival (log-rank test (Mantel-Cox), p=0.019).
Conclusion: Podosomes and their rosettes are driven by Ezrin-Cortactin interaction and this plays a role in pancreatic cancer invasion.