In many physiological and disease processes, TGF-β usurps branches of MAP kinase pathways in conjunction to Smads to induce apoptosis and epithelial to mesenchymal transition, but the detailed mechanism of how a MAP kinase cascade is activated by TGF-β receptors is not clear. We report here that TRAF6 is specifically required for the Smad-independent activation of JNK and p38 and its carboxyl TRAF homology domain physically interacts with TGF-β receptors. TGF-β induces K63-linked ubiquitination of TRAF6, and promotes association between TRAF6 and TAK1. Our results indicate that TGF-β activates JNK and p38 through a mechanism similar to that operating in the interleukin-1β/Toll-like receptor pathway.

Controlled protein degradation mediated by ubiquitin/proteasome system (UPS) plays a crucial role in modulating a broad range of cellular responses. Dysregulation of the UPS often accompanies tumorigenesis and progression. Here, we report that Smad ubiquitination regulatory factor 2 (Smurf2), a HECT-domain containing E3 ubiquitin ligase, is upregulated in certain breast cancer tissues and cells. We show that reduction of Smurf2 expression with specific short interfering RNA in metastatic breast cancer cells induces cell rounding and reorganization of the actin cytoskeleton, which are associated with a less motile and invasive phenotype. Overexpression of Smurf2 promotes metastasis in a nude mouse model and increases migration and invasion of breast cancer cells. Moreover, expression of Smurf2CG, an E3 ligasedefective mutant of Smurf2, suppresses the above metastatic behaviors. These results establish an important role for Smurf2 in breast cancer progression and indicate that Smurf2 is a novel regulator of breast cancer cell migration and invasion. [Cancer Res 2009;69(3):735-40]

Higher cyclooxygenase 2 (COX-2) expression is often observed in aggressive colorectal cancers (CRCs). Here, we attempt to examine the association between COX-2 expression in therapy-refractory CRC, how it affects chemosensitivity, and whether, in primary tumors, it is predictive of clinical outcomes. Our results revealed higher COX-2 expression in chemoresistant CRC cells and tumor xenografts. In vitro, the combination of either aspirin or celecoxib with 5-fluorouracil (5-FU) was capable of improving chemosensitivity in chemorefractory CRC cells, but a synergistic effect with 5-FU could only be demonstrated with celecoxib. To examine the potential clinical significance of these observations, in vivo studies were undertaken, which also showed that the greatest tumor regression was achieved in chemoresistant xenografts after chemotherapy in combination with celecoxib, but not aspirin. We also noted that these chemoresistant tumors with higher COX-2 expression had a more aggressive growth rate. Given the dramatic response to a combination of celecoxib + 5-FU, the possibility that celecoxib may modulate chemosensitivity as a result of its ability to inhibit MDR-1 was examined. In addition, assessment of a tissue microarray consisting of 130 cases of CRCs revealed that, in humans, higher COX-2 expression was associated with poorer survival with a 68% increased risk of mortality, indicating that COX-2 expression is a marker of poor clinical outcome. The findings of this study point to a potential benefit of combining COX-2 inhibitors with current regimens to achieve better response in the treatment of therapy-refractory CRC and in using COX-2 expression as a prognostic marker to help identify individuals who would benefit the greatest from closer follow-up and more aggressive therapy.