Abstract
p53 protein, activated and stabilized by posttranslational modifications, performs its major functions by inducing DNA repair, cell-cycle arrest, or apoptosis through transcriptional activation. Here, we determined the ability of p53 protein stabilized via proteasome inhibition to perform similar functions as p53 induced by stresses such as DNA damage. Treating mice with the proteasome inhibitor bortezomib stabilized p53 in stem/progenitor cells of the intestine and stomach, in other proliferating tissues, and in intestinal tumors. Robust basal p53 mRNA levels were observed in the same compartments where p53 was stabilized. Spatial activation of p53 target genes in response to bortezomib in the small intestine demonstrated that CDKN1A and BAX were upregulated in the proliferative crypts but not in the differentiated villi of the small intestine; PUMA was specifically activated at the crypt base of p53 wild-type mice. Thus, cellular context determines the p53 transcriptional target selection. p53-dependent apoptosis was induced in Lgr5-expressing stem cells of the small intestine and high p53 transcriptional activity and apoptosis was induced in intestinal adenomas and in xenograft tumors. Bortezomib inhibited the growth of intestinal adenomas and xenograft tumors with wild-type p53, indicating the importance of p53 in the response to proteasome inhibitors in tissue homeostasis and in cancer therapy. SIGNIFICANCE: These findings show that bortezomib is less active in p53-defective tumors, yet its success in treating multiple myeloma suggests its use can be extended to p53-proficient solid tumors.
Highlights
Keeping p53 protein levels low in normal cells is necessary for tissue homeostasis and this is maintained by the E3 ubiquitin ligase activity of Mdm2
Coexpression of p53/Sox9 and p53/Lgr5 was www.aacrjournals.org further detected by Co-ISH in nonstressed APCMin/þ mice (Fig. 6E). These results indicate that bortezomib treatment stabilizes p53 in Wnt-driven intestinal adenomas in a similar fashion to that observed in intestinal crypts
Proteasome inhibition by bortezomib resulted in stabilized p53 in normal and tumor cells, facilitating for the first time, the detection of abundant wild-type p53 in vivo independent of stress or DNA damage stimuli
Summary
Keeping p53 protein levels low in normal cells is necessary for tissue homeostasis and this is maintained by the E3 ubiquitin ligase activity of Mdm. Contradictory observations from independent studies have precluded a formal conclusion as to whether p53 contributes to bortezomib-induced apoptosis and whether the stabilization of p53 by proteasome inhibitors induces p53 transcriptional activation. Because wild-type p53 is www.aacrjournals.org known to be highly regulated by ubiquitination and proteasomal degradation in tissue culture cells, we reasoned that this detection failure could be due to the short half-life of the protein in vivo and that we might be able to detect expression of the wild-type p53 protein in proliferative tissues of mice treated with bortezomib. We have investigated the stabilization of p53 and its transcriptional activation by bortezomib in fast renewing normal tissues and tumors, with a major focus on gastrointestinal organs and intestinal adenomas. We successfully documented the stabilization and activation of p53 by bortezomib in discrete epithelial compartments of the small intestine: differentiated cells in the villi, rapidly proliferating cells in the transit-amplifying zone, stem cells at the crypt base, and tumor cells in intestinal adenomas
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