Fig. 7
Timeless activates β-catenin through the Myosin-9/β-catenin pathway. Timeless overexpressing HCT116 and RKO cells were transfected with Myosin-9 specific si-MYH9 (pcDNA-Tim + si-MYH9) or control si-NC (pcDNA-Tim + si-NC). (a) Expression levels of Timeless and Myosin-9 in HCT116 and RKO cells were measured by western blotting. CCK8 (b) and colony-forming growth assays (c) were used to determine the proliferation rate of the indicated CRC cells. Transwell assays were performed to test the migration (d) and invasion (e) of the indicated CRC cells. (f) EMT-associated proteins in the indicated CRC cells were measured by western blotting. Expression level of total β-catenin (g) and subcellular β-catenin (h) in Myosin-9 knockdown cells were detected with Western blot. (i) Expression level of β-catenin was detected in CRC cells with Timeless overexpression and Myosin-9 knockdown. (j) Immunoprecipitation (IP) assay was performed to detect interactions between Myosin-9 and β-catenin. (k) Immunoprecipitation (IP) with anti-Myosin-9 was performed to detect the level of β-catenin in the cells with Timeless knockdown. Error bars indicate the means ± SD. *P < 0.05, **P < 0.01