A mechanosensitive RNAi machinery regulates pro-tumorigenic transformation in the colon

Background: Up to 90% of all cancers are of epithelial origin, such as the cancers of the gastrointestinal tract. A hallmark of the epithelial tissue is that cells that comprise it form a barrier that depends on the integrity of the adherens junctions (AJs), an essential cell-cell adhesion complex. However, a common observation in cancer is that this barrier and the AJs are compromised, promoting cancer progression. We have particularly discovered mislocalization of the AJ and barrier component PLEKHA7 in colon cancer, resulting in pro-tumorigenic cell behavior, caused by dysregulation of a PLEKHA7-associated RNA interference (RNAi) machinery and oncogene upregulation. However, the reasons for this PLEKHA7-RNAi disruption in colon tumors are unclear. To investigate what causes this disruption, we will perturb the extracellular matrix (ECM), which is commonly found to increase in stiffness as cancer progresses, potentially promoting tumor behavior. Our hypothesis is that mechanical changes that occur during cancer progression cause disruption of the PLEKHA7-RNAi machinery. Methods: To test this hypothesis, we have generated several substrates of polyacrylamide gels and functionalized them with collagen to simulate the ranges of elastic moduli seen in normal and diseased colonic tissues. We will be plating non-malignant colon Caco2 cells on these substrates to investigate how ECM stiffness impacts localization of PLEKHA7 and RNAi components at AJs, as examined by immunofluorescence and confocal microscopy. Results and Conclusions: Currently, our results show that increasing ECM stiffness disrupts formation of this AJ complex, suggesting that the elastic modulus of the ECM indeed impacts PLEKHA7-RNAi function and epithelial integrity. We will be conducting further analyses to delineate the exact properties of the elastic moduli of the ECM that result in PLEKHA7-RNAi disruption and the extent that this ECM-PLEKHA7-RNAi crosstalk contributes to disease progression.