FAM29A interacts with NEDD1. (A) Myc-NEDD1 was cotransfected with GFP-FAM29A or GFP into HeLa cells. Lysates of transfected HeLa cells were immunoprecipitated (IP) with anti-GFP antibodies or with nonspecific IgG followed by Western blotting. (B) HeLa S3 cells were synchronized at prometaphase by a thymidine-nocodazole treatment (TN0). Lysates from asynchronous cells (AS) or prometaphase cells were immunoprecipitated with anti-FAM29A antibodies or with nonspecific IgG. The immunoprecipitates were treated with or without λ-phosphatase followed by Western blotting. (C and D) HeLa S3 cells were synchronized at prometaphase by a thymidine-nocodazole treatment (TN) (C) or at the G1/S boundary by a double-thymidine treatment (TT) (D). Cells were released and harvested at the indicated times (hours). Cell cycle profile was determined by FACS. Lysates and the anti-FAM29A immunoprecipitates were analyzed by Western blotting. In B–D, arrowheads point to nonspecific cross-reacting proteins on Western blot. “ap” (in C), apoptotic cells.