Inactivation of p53 the master regulator of cellular stress and damage

Inactivation of p53 the master regulator of cellular stress and damage signals often allows cells that should die or senesce to live. knockout Eμ-transgenic mice. Moreover p53 loss in transformed B-cells did not confer protection from apoptosis as deletion in established deletion MifaMurtide retained at least one allele of deletion in lymphomas reduced tumor burden and prolonged survival. Therefore inactivation of is insufficient to allow untransformed B-cells and B-cell lymphomas to survive without or reduced DICER expression or enzymatic activity is reported in multiple solid organ tumors (6 7 8 9 10 11 12 13 14 15 16 Mouse models revealed Dicer is a haploinsufficient tumor suppressor in soft tissue sarcoma MifaMurtide lung adenocarcinoma and retinoblastoma (17 18 In contrast we showed heterozygosity had no effect on the rate of B-cell lymphoma development (19). Therefore differences in the requirements for Dicer and the effects of reduced Dicer expression in different tissues remain unresolved. The p53 tumor suppressor which induces apoptosis or cell cycle arrest upon cellular stresses (20) responds to defects in miRNA biogenesis and therefore may be required to signal problems in this pathway. Specifically in untransformed murine embryonic fibroblasts (MEFs) deletion of leads to p53 activation and premature senescence which is delayed with loss of (21). We previously detected an increased frequency of inactivation in lymphomas in a mouse model of Myc-induced B-cell lymphoma (Eμ-alleles suggesting a connection between activation and deletion in B-cells (19). Moreover data from three groups including our own showed expression of Cre in mice in B-cell progenitors or mature B-cells results in B-cell apoptosis (19 22 23 This apoptosis was partially rescued by overexpressing the anti-apoptotic Bcl-2 protein or reducing the pro-apoptotic Bim protein (22). Although deletion (23) deletion was synthetically lethal in Dicer and Rb deficient retinal progenitor cells (24). Therefore the role of p53 in monitoring defects in miRNA biogenesis and cell survival in the context of a deficiency remains unclear. Using mouse models we determined the contribution of p53 to B-cell survival and lymphoma development with loss of Dicer. A deficiency did not rescue the defect in B-cell development the reduction in B-cell survival or the delay in Myc-induced lymphomagenesis upon deletion. It did restore the B-cell lymphoma phenotype. However none of the lymphomas that emerged had deleted both alleles of underwent apoptosis when was deleted significantly extending survival in mouse models. Thus p53 loss is insufficient to allow survival and growth of B-cells and B-cell lymphomas MifaMurtide in the absence of Dicer and thus targeting Dicer may have therapeutic potential for treating B-cell lymphomas. MifaMurtide Materials and Methods Mice C57Bl/6 Eμ-(25) and CD19-(26) transgenic mice mice from Dr. Steve Jones (21) and mice from Dr. Guillermina Lozano (27) were intercrossed to obtain the mice needed for this study. Littermates were used in all analyses. For experiments with nude mice 1.5 or 0.5×106 deleted lymphoma cells expressing a tamoxifen-inducible form of Cre (CreERT2) were injected (subcutaneous or intravenous respectively) into 6-week-old female mice (Harlan labs). Tamoxifen (2 mg) or corn oil (vehicle control) was injected (intraperitoneal) once daily for 3 days starting the day of lymphoma injection for two cohorts (one IL18RAP subcutaneous and one tail vein injected cohort) or after lymphomas were 90-150mm3 for a second subcutaneous cohort. Subcutaneous tumors were measured with calipers and tumor volume calculated. Blood was collected for flow cytometric and microscopic analyses from the mice where lymphoma was injected into the tail vein. Mice were humanely sacrificed prior to lymphoma development or for survival studies at humane endpoints and tumors/tissues MifaMurtide were harvested and analyzed. Log-rank tests determined statistical significance for survival. All studies were in accordance with state and federal guidelines and were approved by the Vanderbilt Institutional Animal Care and Use Committee. Western and Southern blotting Whole cell protein lysates from B-cell lymphomas and pre-B cells were generated and Western blotted as previously described (28). Antibodies against p19Arf (GeneTex) p53 (Ab-7; Calbiochem) Mdm2 (C-18; Santa Cruz) Cre.