Fig. 6.

Feline cancer stem cells lack activation of the p53 DNA damage pathway in response to doxorubicin and ionising radiation. (A) Dissociated mammospheres and parental adherent cells were treated with 10 μM doxorubicin or dimethyl sulfoxide (DMSO), cells were harvested over the indicated time course and expression of p53 pathway related proteins was assessed. (B) Dissociated mammospheres and parental adherent cells were treated with 5 Gy ionising radiation, cells were harvested over the indicated time course and expression of proteins related to the p53 pathway was assessed. Dissociated mammospheres and adherent cells were incubated with 2.5 μM doxorubicin (Dox) or DMSO. Proteins were extracted according to their subcellular localisation: F1, cytosolic; F2, membranes/organelles; F3, nucleus; F4, nucleus; F5, cytoskeleton. Proteins from each fraction were resolved by SDS–PAGE and analysed by immunoblotting for p53; 30 μg was loaded per lane. Coomassie (CM) staining confirmed that protein expression profiles from each fraction were distinct; 5 μg was loaded per lane (C).