Histone h3k27me3

The following antibodies were used at the indicated dilutions: PME‐1, Santa Cruz Biotechnology (Dallas, TX, USA) sc‐20086 (H‐226), Western blotting 1 : 1000; PME‐1, Santa Cruz Biotechnology sc‐25278 (B‐12), Immunohistochemistry 1 : 1000, Immunofluorescence 1 : 100; cleaved PARP‐1, Abcam (Cambridge, UK) ab32064 [E51], Western blotting 1 : 1000; GAPDH, HyTest 5G4‐6C5, Western blotting 1 : 5000; c‐MYC, Abcam ab32072 [Y69], Western blotting 1 : 1000; p‐Myc S62, Abcam ab78318, Western blotting 1 : 1000; AKT1/2/3, Cell Signaling Technology #9272, Western blotting 1 : 2000; p‐AKT S473, Cell Signaling Technology #4060, Western blotting 1 : 1000; Lamin‐A/C, Santa Cruz Biotechnology sc‐7292 (636), Immunofluorescence 1 : 250; Lamin‐A/C, Santa Cruz Biotechnology sc‐6215 (N‐18), Western blotting 1 : 1000; p‐Lamin‐A/C S392, Abcam ab58528, Western blotting 1 : 5000; EEA1, Santa Cruz Biotechnology sc‐137130 [G4], Immunofluorescence 1 : 100; p‐FAK Y397, Cell Signaling Technology #8556, Immunofluorescence 1 : 100; Histone H3K9me3, Cell Signaling Technology #13969 [D4W1U], Immunofluorescence 1 : 500; Histone H3K27me3, Cell Signaling Technology #9733 [C36B11], Immunofluorescence 1 : 500; PPP2R2A, Cell Signaling Technology #5689, Western blotting 1 : 1000.

Total cell lysates were prepared in a buffer [50 mM Tris‐HCl (pH 7.5), 150 mM NaCl, 1% IGEPAL, 0.1% SDS, and 0.5% sodium deoxycholate] supplemented with a mixture of protease and phosphatase inhibitors. Protein concentrations were measured using the bicinchoninic acid protein assay reagent kit (Pierce Chemical Co). Equal amounts (20ug) of protein lysates from various samples were used for SDS–PAGE analysis followed by immunoblotting. Antibodies to cyclin B1 (12231), cyclin A2 (4656), cyclin D3 (2936), vinculin (13901), Eg5 (14404), Histone H3 K27 Me3 (9733), Histone H2A K119 Ub1 (8240), phospho‐Histone H3 (Ser 10) (3377), DYKDDDDK (FLAG Tag) (14793), BubR1 (4116), CBX8 (14696), PARP‐1 (9542), α‐Tubulin (2144), Histone H2A (12349), phospho‐Histone H2Ax (2577) and β‐Actin (4970) were purchased from Cell Signaling Technology. Anti‐GST (Z‐5), Bub3 (H‐100), Histone H3 (FL‐136) and p55 CDC (H‐175) (Cdc20) antibody were purchased from Santa Cruz biotechnology. Antibodies to MAD2 (A300‐301A) and PTEN (A300‐700A) were obtained from Bethyl laboratories. Specific signals on immunoblots (polyvinylidene difluoride) were visualized using enhanced chemiluminescence (Super‐Signal, Pierce Chemical Co.).