FIGURE 4.

DOC2B expression changes cell morphology, induces remodeling of cytoskeletons, inhibits cell division, and increases intracellular calcium flux. A, representative cell morphology of DOC2B expressing and control cells in SiHa and HeLa cells, respectively (×40 magnification). B, representative images of actin staining showing rearrangement of actin fibers leading to increased cell to cell adhesion and decreased lamellipodia (indicated by arrows) in DOC2B expressing cells when compared with control cells (×400 magnifications). C, representative anoikis analysis for empty vector- and DOC2B-transfected SiHa and HeLa cells. Cells were cultured on poly-HEMA-coated tissue culture plates and cell death was analyzed by measuring the sub-G₁ population of cells by propidium iodide staining and FACS analysis. Ectopic expression of DOC2B showed a significantly higher sub-G₁ population of 18.66 versus 36.33% and 24.47 versus 48.96% between DOC2B and empty vector-transfected cells in both SiHa and HeLa cells, respectively. D, quantification of DNA content and cell cycle phase distribution in empty vector- and DOC2B-transfected SiHa cells at different time points as analyzed by a BrdU pulse-chase experiment. Ectopic expression of the DOC2B gene resulted in a significant cell cycle arrest at G₀/G₁ and S phases of the cell cycle. E, representative figures showing an increase in intracellular Ca²⁺ flux when compared with control cells. There was a significant increase in intracellular Ca²⁺ upon ectopic expression of DOC2B. Values were the median fluorescence intensities ± S.D. of at least three independent experiments performed in duplicates. The bar graph represents mean ± S.D. of triplicate experiments performed in duplicates. *, p < 0.05 by independent Student's t test. p value <0.05 was considered statistically significant.