Parvovirus B19 NS1 protein induces cell cycle arrest at G2-phase by activating the ATR-CDC25C-CDK1 pathway
Fig 1
Putative Transactivation Domain 2 (TAD2) is essential for NS1-induced G2-phase cycle arrest.
(A) Western blot analysis. NS1-S1 and NS1mTAD2-S1 cell lines were treated with different concentrations of Dox (1, 5, or 10 μg/ml) to induce expression of NS1 or NS1mTAD2, respectively. After 72 h, cell lysates were prepared and probed for NS1 and β-actin expression using anti-strep and anti-β-actin antibodies, respectively. (B) Cell cycle analysis. NS1-S1 and NS1mTAD2-S1 cells were treated with Dox (Dox+; 5 μg/ml) or not (Dox-) for 72 h and then incubated with BrdU, followed by treatment with 1N HCl. Treated cells were stained with an anti-BrdU antibody, and secondary antibody, and DAPI prior to flow cytometry. The numbers in each histogram are percentages of the cell populations with all BrdU-positive (S-phase), and BrdU-negative cell populations of 2N and 4N DNA content (G1- and G2-phase, respectively). (C) Statistical analysis. The percentage of cells at G1, S, and G2 are depicted in color. The percentages of the cells at G2-phase are shown in numbers, and compared in pairs as shown. **P<0.01.