PINT modulates cell proliferation and apoptosis. (A) Inhibition of Pint. Pint levels were detected by quantitative real time (RT-qPCR) in p53-restored doxorubicin-treated p53LSL/LSL MEFs 36 hours after transfection with two Pint-specific anti-sense oligonucleotides (ASOs) (ASO1 and ASO2), two control ASOs (control ASO -1 and -2), or a blank (PBS) control, and 12 hours of doxorubicin treatment. Values normalized to Gapdh and are the mean ± SD of three replicates. (B) Pint positively regulates cell proliferation. Relative number of p53-restored p53LSL/LSL mouse embryonic fibroblasts (MEFs) transfected with ASOs for Pint inhibition, and treated with doxorubicin from 24 h post-transfection. Cell numbers are determined by MTS assay. Values are mean ± SD of three replicates. (C) Overexpression of Pint. Pint levels where measured like in (A) in p53-restored doxorubicin-treated p53LSL/LSL MEFs 36 hours after transfection and 12 hours of doxorubicin treatment with Pint A isoform expressing plasmid or an empty plasmid as control. (D) Pint positively regulates cell proliferation. Cells were transfected as in (C) and treated with doxorubicin from 24 hours post-transfection. (E,F). Negative effect of Pint on apoptosis induction. Apoptosis levels were determined by quantification of caspase 3/7 levels after (E) inhibition or (F) overexpression of Pint in p53-restored p53LSL/LSL MEFs treated with doxorubicin. Values are the mean ± SD of three replicates. (G,H). Effect of Pint on cell cycle regulation. Relative cell numbers in each cell cycle phase were determined by fluorescence-activated cell sorting (FACS) of bromodeoxyuridine (BrdU) incorporation and propidium iodide (PI) staining of p53-restored p53LSL/LSL MEFs treated as in (A) or (C). Percentages of cells in each phase are represented and values are the mean ± SD of three replicates.