ABSTRACT
BACKGROUND: We investigated the association between the increased eukaryotic translation initiation factor 4E (eIF4E) level and MDM2 overexpression in the esophageal cancer tissue and cells. METHODS: This was a retrospective study of specimens from esophageal cancer patients treated over a 5-year period in a Taiwan university hospital. The predictor variable was eIF4E level in esophageal tumors and CE48T/VGH and TE6 esophageal carcinoma cell lines. The main outcome variable was MDM2 overexpression. Appropriate descriptive and univariate statistics were computed, and a P value of <0.05 was considered statistically significant. RESULTS: There were two study sample groups. Immunohistochemistry analyses of the first sample group (51 esophageal tumors) revealed that 19 specimens demonstrated MDM2 elevation and 20 specimens had eIF4E overexpression. eIF4E elevation was evidenced by accumulation of the protein in the cytoplasm. There was a significant association between the eIF4E and MDM2 expression (P < 0.001). Western blot analysis and semiquantitative reverse transcriptase-polymerase chain reaction of the second specimen group (20 pairs of tumors and normal tissues) revealed the co-elevation of MDM2 and eIF4E (P = 0.008). There was no increased mdm2 transcript in most of the specimens. Without significant alterations in the mdm2 mRNA level and subcellular distribution, MDM2 protein was upregulated in CE48T/VGH cultured cells expressing ectopic eIF4E. Conversely, reduction of eIF4E by specific siRNA enabled TE6 cells synthesizing reduced amounts of MDM2. CONCLUSIONS: Our findings indicate that MDM2 protein levels are strongly associated with and regulated by eIF4E in a posttranscriptional mechanism in esophageal cancer.
Subject(s)
Carcinoma, Squamous Cell/genetics , Esophageal Neoplasms/genetics , Esophagus/metabolism , Eukaryotic Initiation Factor-4E/genetics , Proto-Oncogene Proteins c-mdm2/genetics , Adult , Aged , Aged, 80 and over , Blotting, Western , Carcinoma, Squamous Cell/metabolism , Carcinoma, Squamous Cell/secondary , Case-Control Studies , Esophageal Neoplasms/metabolism , Esophageal Neoplasms/pathology , Esophagus/pathology , Eukaryotic Initiation Factor-4E/metabolism , Female , Humans , Immunoenzyme Techniques , Lymphatic Metastasis , Male , Middle Aged , Neoplasm Invasiveness , Prognosis , Proto-Oncogene Proteins c-mdm2/antagonists & inhibitors , Proto-Oncogene Proteins c-mdm2/metabolism , RNA, Messenger/genetics , RNA, Small Interfering/genetics , Retrospective Studies , Reverse Transcriptase Polymerase Chain Reaction , Tumor Cells, CulturedABSTRACT
Rapamycin, a potential anti-cancer agent, modulates activity of various factors functioning in translation, including eIF4E, an initiation factor selectively regulating expression of a subset of cellular transcripts. We show here that rapamycin suppresses levels of the p53-regulator MDM2 by translational inhibition without affecting mdm2 mRNA expression or protein stability. Rapamycin inhibits translation of mdm2 mRNA from the constitutive P1 promoter, which contains two upstream ORFs (uORFs) in the 5'UTR. Suppression is accompanied by increased hypo-phosphorylation of 4EBP-1, an inhibitory eIF4E binding protein. Ectopic expression of eIF4E abrogates rapamycin-mediated MDM2 inhibition, suggesting that eIF4E is crucial in modulating MDM2 expression in rapamycin-treated cells. Rapamycin administration also results in elevated PUMA expression and PARP cleavage, which is reproduced by siRNA knockdown of eIF4E or MDM2, suggesting that MDM2 suppression by rapamycin stimulates p53-mediated apoptosis. Together, our results define translational regulation of MDM2 expression by eIF4E and provide a molecular mechanism underlying rapamycin-induced p53-dependent apoptosis.
