Ptch1

Tissues were snap-frozen in liquid nitrogen and stored at −80 °C. Pulverized frozen tumor samples were lysed in radioimmunoprecipitation assay (RIPA) buffer and the total proteins were extracted, separated, and transferred using standard procedures. Antibodies were purchased from BD Transduction Laboratories: SMARCB1 (1/500, 612110); Cell Signaling Technology: Akt (1/2000, #9272), P-Akt (Ser473) (1/2000, #4060), CDK4 (1/2000, #2906), CDK6 (1/2000, #3136), Cyclin D1 (1/3000, #2926), merlin (1/1000, #6995), p27 (1/1000, #2552), P-Rb (1/1000, #8516), S6 (1/1000, #2317), P-S6 (Ser 235/236) (1/4000, #4858), 4E-BP1 (1/2000, #9452); Novus Biologicals: GAPDH (1/5000, NB300-221), PTCH1 (1/1000, MAB41051), RB1 (1/50, NB120-3077), SHH (1/500, AF464); R&D Systems: GLI-1 (1/1000, AF3455); Santa Cruz Biotechnology: p16 (1/3000, sc-1207) and Thermo Scientific: p21^WAF1 (1/1000, MS-387-P0). All uncropped scans of western blots are provided in Supplementary Fig. 7.

Traditional western blots were performed using standard protocols. Primary antibodies for B-Actin(1:1000, Cat #8457S, Cell Signaling Technologies), Vinculin(1:30000, Cat #V9264, Sigma Aldrich), PTCH1(1:500,Cat#MAB41051, R&D systems) and APC(1:500, Cat #15270, Abcam) were used. Knock out HSC1λ cell line genotypes were confirmed with capillary electrophoresis-western blot using a WES capillary electrophoresis device (ProteinSimple, San Jose, CA, USA) according to the manufacturer’s instructions. Primary antibodies against B-Actin (Cell Signaling Technologies #8457S, 1:10), PTCH1 (Cell Signaling Technologies #2468, 1:10), GLI1 (Cell Signaling Technologies #3538, 1:10).