Regulation of mdm2 mRNA expression in human breast tumor-derived GI-101A cells.
The MDM2 oncoprotein (p90) binds to p53 and inhibits its function. Here, the expression of mdm2 mRNA subsequent to phorbol 12,13-dibutyrate (PDB) or diethylstilbestrol (DES) treatment was analyzed in human breast tumor-derived GI-101A cell line. Expression of mdm2 mRNA was detected in rapidly growing GI-101A cells and that was similar to the expression seen in HL-60 cells. On the other hand PC12 (rat adrenal pheochromocytoma cells) did not show any mdm2 expression. GI-101A cells were treated with varying concentrations of DES or PDB, and mdm2 mRNA levels were determined by RT-PCR analysis. The RT-PCR results clearly showed that mdm2 mRNA expression was increased with increasing concentrations of PDB and DES treatments. To determine the specificity of the effects produced by DES and PDB the cells were treated with estrogen receptor antagonist tamoxifen and protein kinase C (PKC) specific inhibitor chelerythrine. Tamoxifen and chelerythrine co-treatments inhibited DES and PDB stimulated increases of mdm2 transcription respectively, in GI-101A cells. In an attempt to determine the upstream signaling pathway, the effects of PDB or DES on the mitogen activated protein kinase (MAPK) levels were determined by western blot analysis in the presence and absence of PD098059, a specific inhibitor of mitogen activated protein kinase kinase (MAPKK). The phospho-MAPK (p44/42) levels, an activated form of MAPK, increased in DES and PDB stimulated cells whereas PD098059 treatment inhibited this increase. Our data implicate MAPK as an upstream regulator of mdm2 expression and help to speculate on the intracellular regulation of mdm2 expression.
Zell
Jason A
JA
College of Osteopathic Medicine, Health Professions Division, Nova Southeastern University, Ft. Lauderdale, FL 33328, USA.
Ramakrishnan
Ramugounder
R
Rathinavelu
Appu
A
D000470:Alkaloids; D000818:Animals; D001483:Base Sequence; D053119:Benzophenanthridines; D001943:Breast Neoplasms / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D017931:DNA Primers; D004054:Diethylstilbestrol / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009687:Nuclear Proteins; D016716:PC12 Cells; D010617:Phenanthridines / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D051736:Proto-Oncogene Proteins c-mdm2; D012333:RNA, Messenger / Q000235:genetics*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013629:Tamoxifen / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000470:Alkaloids; D053119:Benzophenanthridines; D017931:DNA Primers; D005419:Flavonoids; D009687:Nuclear Proteins; D010617:Phenanthridines; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D013629:Tamoxifen; D015240:Phorbol 12,13-Dibutyrate; D004054:Diethylstilbestrol; C016299:chelerythrine; C497206:MDM2 protein, human; C497208:Mdm2 protein, rat; D051736:Proto-Oncogene Proteins c-mdm2; D020928:Mitogen-Activated Protein Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one