Mdr1b facilitates p53-mediated cell death and p53 is required for Mdr1b upregulation in vivo

The mdr1b gene is thought to be a "stress-responsive" gene, however it is u nknown if this gene is regulated by p53 in the whole animal. Moreover, it i s unknown if overexpression of mdr1b affects cell survival. The dependence of mdr1b upon p53 for upregulation was evaluated in p53 knockout mice. Wild -type (wt) or p53-/- mice were treated singly or in combination with gamma irradiation (IR) and/or the potent DNA damaging agent, diethylnitrosoamine (DEN), Both IR and DEN induced mdr1b in wild-type animals, but not in the p 53-/- mice. IR also upregulated endogenous mdr lb in the H35 liver cell lin e, and the mdr lb promoter was activated by IR and activation correlated wi th p53 levels; moreover activation required an intact p53 binding site. Col ony survival studies revealed that co-transfection of both mdr1b and p53 dr amatically reduced colony numbers compared to cells transfected with either p53 or mdr1b alone and cells microinjected with both mdr1b and p53 had a m ore dramatic loss in viability compared to cells injected with either expre ssion vector alone. Further studies using acridine orange and ethidium brom ide to measure apoptosis revealed that mdr1b caused apoptosis and this was enhanced by p53, however the increased apoptosis required a functional p53 transactivation domain. These studies indicate that mdr1b is a downstream t arget of p53 in the whole animal and expression of mdr1b facilitates p53-me diated cell death.