Anti cyclin d1 m20

Protein extract preparation^{62 (link)}, immunoprecipitation assay^{63 (link)}, immunoblotting assays^{64 (link)}, and sucrose gradient for rafts isolation^{45 (link)} were performed as described elsewhere. Primary antibodies were as follows: anti-Notch3 (Catalog number 2889), anti-Notch1 (Catalog number 2421), anti-activated Notch1 (N1_Val1744), anti-EGFR D38B1 (Catalog number 4267S), anti-phospho-EGFR Y1173 (53A5-Catalog number 4407S), and anti-phospho-EGFR Y1068 (Catalog number 2234S), all from Cell Signaling (Danvers, MA, USA); anti-α-tubulin (Catalog number sc-8035), anti-Lamin B M20 (Catalog number sc-6217), anti-p27 C19 (Catalog number sc-528), anti-cyclin D1 M20 (Catalog number sc-718), anti-cyclin D3 C16 (Catalog number sc-182), anti-Notch1 L18 (N1_EC) (Catalog number sc-23299), anti-transferrin H65 (Catalog number sc-21011), and anti-PTPH1 (Catalog number sc-515181), all from Santa Cruz Biotechnology; anti-β-actin (Catalog number A5441) and anti-cholera toxin B subunit peroxidase conjugate (GM1, Catalog number C3741) from Sigma-Aldrich; and anti-Notch3 5E1 (N3_EC) antibody was kindly provided by Professor A Joutel⁶⁵ .

Western blotting was performed as previously described [37 (link)]. The primary antibodies used were as follows: anti-Myc-tag antibody (#2276; 1:1000), anti-peIF4E Ser-209 (#9741; 1:1000), anti-Mnk1 C4C1 (#2195; 1:1000), anti-eIF4G (#2498; 1:1000), anti-YB1 (#4202; 1:1000), anti-mTOR 7C10 (#2983; 1:1000), anti-β-catenin (#9562; 1:1000; all from Cell Signaling); anti-EIF4ENIF1 (three different antibodies: 1:500; Sigma; 1:1000 Cell Signaling; 1:1000 Abnova); anti-Cyclin D1 M-20 (1:1000; Santa Cruz Biotechnology); anti-MCL1 Y37 (1:500), anti-Ago2 (1:500), anti-GAPDH 6C5 (1:2000; all from Abcam); and anti-HuR (1:500; Millipore). Anti-actin CP01 (1:500; Calbiochem, Darmstadt, Germany) was used as a loading control. The secondary antibodies used were donkey anti-rabbit IgG-HRP (NA9340; 1:2000) and donkey anti-mouse IgG-HRP (NA9340; 1:2000; both from Amersham Pharma-Biotech, Uppsala, Sweden). Bound antibodies were visualized with an enhanced chemiluminescence detection kit (Amersham Pharma-Biotech).