A novel RIP1-mediated canonical WNT signaling pathway that promotes colorectal cancer metastasis via ß -catenin stabilization-induced EMT.

ABSTRACT

RIP1 (receptor-interacting protein kinase 1) is an important component of TNF-α signaling that contributes to various pathological effects. Here, we revealed new potential roles of RIP1 in controlling WNT/ß-catenin canonical signaling to enhance metastasis of colorectal cancer (CRC). First, we showed that WNT3A treatment sequentially increased the expression of RIP1 and ß-catenin. Immunohistochemical analyses of human CRC tissue arrays consisting of normal, primary, and metastatic cancers indicated that elevated RIP1 expression might be related to ß-catenin expression, carcinogenesis, and metastasis. Intravenous injection of RIP1 over-expressed CRC cells into mice has demonstrated that RIP1 may promote metastasis. Immunoprecipitation (IP) results indicated that WNT3A treatment induces direct binding between RIP1 and ß-catenin, and that this stabilizes the ß-catenin protein in a manner that depends on the regulation of RIP1 ubiquitination via downregulation of the E3 ligase, cIAP1/2. Elimination of cIAP1/2 expression and inhibition of its ubiquitinase activity enhance WNT3A-induced RIP1 and ß-catenin protein expression and binding, which stimulates endothelial-mesenchymal transition (EMT) induction to enhance the migration and invasion of CRC cells in vitro. The results of the in vitro binding assay and IP of exogenous RIP1-containing CRC cells additionally verified the direct binding of RIP1 and ß-catenin. RIP1 expression can destroy the ß-catenin-ß-TrCP complex. Taken together, these results suggest a novel EMT-enhancing role of RIP1 in the WNT pathway and suggest a new canonical WNT3A-RIP1-ß-catenin pathway that contributes to CRC malignancy by promoting EMT.