Absence of synergism of genotoxins and chromatin modifiers effects on p19 induction.

<p><b>A.</b> HEK-293 cells were exposed to 40 J/m² UV or 50 ng/ml neocarzinostatin and incubated in the presence or in the absence of 100 µM chloroquine, or 200 nM TSA or hypotonic medium. After 4 h, cells were harvested and subjected to northern blot analysis using a ³²P-labelled probe specific for human p19 mRNA and reprobed for E2F1 and β-tubulin mRNA. <b>B.</b> HEK-293 cells were transfected with increasing levels of an expression vector encoding E2F1 gene, harvested after 24 h, and p19 and E2F1 expression assessed by northern blot. In parallel cells were incubated with 100 µM chloroquine or 200 nM trichostatin A or hypotonic medium or exposed to 40 J/m² UV as indicated. After 4 h p19 and E2F1 expression was determined by northern blot. In (<b>A</b>) and (<b>B</b>) each figure shows a representative autoradiograph of three independent experiments with similar results. Densitometric analysis of p19 and E2F1 are represented in the lower panels. Bars represent the mean ± S.D. of three experiments. Student’s <i>t</i>-test was used to compare treated and non-treated samples (* p<0.05, at least). <b>C.</b> HEK-293 cells, transiently cotransfected with 4 µg of p19CAT and 5 µg pCEFL-β-galactosidase, were exposed to 40 J/m² UV or treated with 50 ng/ml NCS and incubated in the presence or in the absence of 100 µM chloroquine or 200 nM TSA or hypotonic medium. After 24 h cells were harvested and CAT activity was determined as described. Results are expressed as relative CAT activity with respect to basal value of p19CAT which was set to 100. Bars represent the mean ± S.D. of three independent experiments performed in quadruplicate. β-tubulin (β-tub), chloroquine (chlo), neocarzinostatin (NCS), hypotonic medium (hypo).