Figure 5.

NEDD4 serves as an E3 ubiquitin ligase targeting FOXA1 degradation. A, potential E3 ubiquitin ligases that can bind to E3 predicted on ubibrowser; B, binding relationship between Lovo and Caco-2 examined by IP in Lovo and Caco-2 cells; C, subcellular localization of NEDD4 and FOXA1 examined by immunofluorescence double staining; D, NEDD4 and Flag-FOXA1 vector were co-transfected in 293 T cells, and the binding relationship between NEDD4 and FOXA1 was further validated by IP using the anti-Flag and Anti-NEDD4 antibodies; E, overexpression of HA-ubiquitin, flag-FOXA1 or NEDD4 was introduced in 293 T cells, and then the ubiquitination level of FOXA1 was validated by IP; F, NEDD4-mediated ubiquitination regulation on FOXA1 validated by an in-vitro ubiquitination assay. At least three independent experiments were performed