Effects of mutation of MSP58 NLSs on p21 gene expression and cell proliferation. A and B. Growth curves of HT1080 and HeLa, stable control (vector); wild type; NLS1 mutation (10A) and NLS2 mutation (5A) MSP58-overexpressing cells. Points, mean value from three independent experiments, each run in triplicate; bars, S.D. MSP58 expression was analyzed using immunoblotting. C. Phase-contrast images and SA-β-gal activity of control vector and MSP58 overexpression HT1080 stable cell lines. Images are all at the same magnification (200×). D. Total cell extracts were analyzed using immunoblotting with the indicated antibodies. E. p21 mRNA expression was analyzed by quantitative RT-PCR using GAPDH levels as the internal control. The expression of p21 in HT1080 control vector cells was defined as 1.0, and other values were normalized accordingly. Columns, mean of three independent experiments; bars, S.D. *, p < 0.05. NS, non-significant.