Fig. 3

Cell-cycle-dependent centrosomal enrichment of GRP75 and its phosphorylation acting on sub-phase distribution. (A) Cyclebase-based predication of expression of GRP75, MPS1, VDAC1 and Arf6 in sub-phases of cell-cycle (dark blue content indexes the mRNA level during cell-cycle). (B) Western blot determined the expression level of those mentioned protein in subcellular fractions of synchronized Skov3. Protein bands were quantified by Image J software and expression ratio (compared to cells without drug-induction, Ctrl, set as 1) in the sub-phase are correspondingly marked below. (C) Schematic GRP75 constructs: wt: wild type. unphospho: unphosphorlation (T62A/S65A). phospho mimic: phosphorlation mimic (T62D/S65D). (D) Lentivirus-stable-transduced Skov3 and Cos7 cells were built based on GRP75-targeting shRNA (GRP75-KD), or over-expression (OE) with constructs as listed above. GRP75 expression level was analyzed by western blotting. (E) Bar graphs show the sub-phase percentage of cell-cycle in GRP75-KD or -OE cells. Data are the average results of three independent experiments