Talpid3

The following primary antibodies were used: acetylated α-tubulin (T7451, diluted 1:2000 for immunofluorescence, Sigma), γ-tubulin (T6557, Sigma), FLAG (F1804, Sigma), HA (H3663, Sigma), CEP83 (HPA038161, Sigma), CEP164 (SAB3500022, Sigma), CEP290 (ab84870, Abcam), TALPID3 (24421-1-AP, Proteintech), ARL13b (17711-1-AP, Proteintech), FBF1 (11531-1-AP, Proteintech), TCTN1 (15004-1-AP, Proteintech), SCLT1 (14875-1-AP, Proteintech), IFT140 (17460-1-AP, Proteintech), β-actin (sc-47778, diluted 1:5000 for western blotting, Santa Cruz), SMO (sc-166685, Santa Cruz), CEP89 (ab204410, Abcam), ANKRD26 (GTX128255, GeneTex), polyglutamylated tubulin (ALX-804-885-C100, diluted 1:2000 for immunofluorescence, Enzo Life Sciences), GLI-3 (AF3690, R&D Systems), ODF2 (H00004957-M01, Abnova), and Polycystin-2 (Baltimore Polycystic Kidney Disease (PKD) Research and Clinical Core Center). The secondary antibodies were goat anti-mouse Alexa Fluor 488/594 or goat anti-rabbit Alexa Fluor 488/594 (1:1000 dilution). Primary antibodies were diluted 1:500 for immunofluorescence and 1:2000 for western blotting experiments unless otherwise specified. Uncropped versions of blots can be found in Supplementary Fig. 9.

HEK293T cells, wild type C3H10T1/2 cells, C3H10T1/2^Fbxw7KO, C3H10T1/2^Nde1KO and C3H10T1/2 ^{Fbxw7KO;Nde1KO} clones were lysed in 1% Triton X-100, 150 mM NaCl, 10 mM Tris-HCl at pH 7.5, 1 mM EGTA, 1 mM EDTA, 10% sucrose, and a protease inhibitor cocktail (Roche Applied Science), phosphatase inhibitor cocktail (PhosSTOP EASYpack, ROCHE) at 4 °C for 30 min. Cell lysates were separated with SDS-PAGE. Antibodies were used against ALP (Thermofisher, 1:200), NDE1 (Proteintech, 1:1000), β-tubulin (Santa Cruz, 1:1000), TALPID3 (Proteintech 1:1000), HA (Santacruz 1:1000), MYC-Tag (Cell Signaling), FLAG (Sigma 1:1000), FBW7 (Bethyl Laboratories. A301‐720A; A301‐721A 1:500), NDEL1 (Abcam, 1:1000), GAPDH (GeneTex, 1:2000), and IFT88 (Proteintech, 1:000). Densitometric quantification was performed with the Licor Image Studio software.