Fig. 1: CDK2 interacts with and phosphorylates NF90.

a, b Flag-NF90 immunoprecipitated with Myc-CDK2. After co-transfection with Flag-NF90 and Myc-CDK2 plasmids, HEK293T cell lysates were immunoprecipitated by either Flag or Myc antibody then subjected to western blot (WB) analysis. c Association between endogenous NF90 and CDK2. Cell lysates were collected for immunoprecipitation with the CDK2 antibody and subjected to WB as shown. d Schematic view of truncates containing different domains of NF90. e GST-pulldown assay to show which domain of NF90 is involved in the interaction with CDK2. Truncates with GST tag were purified and incubated with cell lysate. f NF90-WT and NF90-S382A plasmids were transfected into 293T cells. After 24 h, cells were lysed and NF90-WT and NF90-S382A were immunoprecipitated by M2 beads then subjected to WB. S382-P, antibody specifically recognizes the phosphorylated NF90-Ser382. g Phosphorylation of NF90-Ser382 was inhibited by roscovitine. 24 h after transfection, cells harboring NF90-WT were treated with 20 µM roscovitine for 24 h. IP and WB assays were performed as described above. h Cells were treated with 20 µM or 40 µM roscovitine for 24 h to detect the phosphorylation level of NF90-S382. IP and WB assays were performed as described above. i HeLa cells were arrested at the G1/S boundary twice by a double-thymidine block, then released and harvested at the indicated time points. Cell lysates were subjected to WB. β-actin served as a loading control.