Research Paper Volume 12, Issue 19 pp 19273—19292

USP18 regulates c-Myc expression through Notch1. (A, B) Co-IP showing that endogenous USP18 and c-Myc in PC cells were not directly bound. (C) Mass spectrometry data show that Notch 1 and USP18 are combined. (D and E) Co-IP showing direct binding of endogenous USP18 and c-Myc in pancreatic cancer cells. (F) Co-localization studies of pancreatic cancer cells using an anti-USP18 antibody (1:200, green) and an anti-Notch1 antibody (1:200, red), followed by DAPI nuclear counterstaining (blue). The merged images of USP18 (green) and Notch1 (red) with DAPI (blue) are also shown. (G and I) Protein and mRNA levels of c-Myc assessed by western blotting and qRT-PCR in pancreatic cancer cells transfected with shNotch1 or shNC. (H and J) Protein and mRNA levels of c-Myc assessed by western blotting and qRT-PCR in pancreatic cancer cells treated with Notch1 inhibitor (Tangeretin). (K and L) Protein levels of Notch1 and c-Myc in pancreatic cancer cells transfected with the indicated plasmid or treated with Notch1 inhibitor (Tangeretin) as assessed by western blotting.