14 3 3 cluc

LATS kinase activity was monitored by LATS-BS consisting of two vectors: NLuc-YAP15 (#107610, Addgene) and 14-3-3-CLuc (#107611, Addgene)^{15 (link)}. Lenvatinib-resistant PLC/PRF/5 and Huh7 were transfected with LATS-BS and pRL-CMV Renilla luciferase (Promega) for normalization of transfection efficiency using Lipofectamine 2000. Treatment of 10 µM U0126 or solvent control (DMSO) for 4 hours was done at 48 hour post transfection. The cells were collected and luciferase signal was measured using Dual-luciferase report assay system (#E1910, Promega). Luciferase fold change was calculated according to the method described^{15 (link)}.

The CDK6-WT and CDK6-K43M mutant pLenti.6.2/V5-DEST^TM vectors were kind gifts from Dr. Claudia Scholl (National Center for Tumor Diseases and German Cancer Research Center (DKFZ)). LATS-Biosensor (NLuc-YAP15 (#107610) and 14-3-3-CLuc (#107611)) were obtained from Addgene. Lenvatinib was purchased from Selleckchem (in vitro experiments) and LC Laboratories (in vivo experiments). The CDK4/6 inhibitor, palbociclib, and the MEK inhibitor, U0126, were purchased from LC Laboratories. The YAP inhibitor, CA3, was purchased from Selleckchem. The PROTAC CDK6 degrader, CP-10, was purchased from MedChemExpress, while BSJ-03-123 was purchased from Tocris Bioscience. MG132 was purchased from Calbiochem.