Multiple levels of regulation of selenoprotein biosynthesis revealed from the analysis of human glioma cell lines

To gain a better understanding of the biological consequences of the exposu re of tumor cells to selenium, we evaluated the selenium-dependent response s of two selenoproteins (glutathione peroxidase and the recently characteri zed 15-kDa selenoprotein) in three human glioma cell lines. Protein levels, mRNA levels, and the relative distribution of the two selenocysteine tRNA isoacceptors (designated mcm(5)U and mcm(5)Um) were determined for standard as well as selenium-supplemented conditions. The human malignant glioma ce ll lines D54, U251, and U87 were maintained in normal or selenium-supplemen ted (30 nM sodium selenite) conditions. Northern blot analysis demonstrated only minor increases in steady-state GSHPx-1 mRNA in response to selenium addition. Baseline glutathione peroxidase activity was 10.7 +/- 0.7, 7.6 +/ - 0.7, and 4.1 +/- 0.7 nmol NADPH oxidized/min/mg protein for D54, U251, an d U87, respectively, as determined by the standard coupled spectrophotometr ic assay. Glutathione peroxidase activity increased in a cell line-specific manner to 19.7 +/- 1.4, 15.6 +/- 2.1, and 6.7 +/- 0.5 nmol NADPH oxidized/ min/mg protein, respectively, as did a proportional increase in cellular re sistance to H2O2, in response to added selenium. The 15-kDa selenoprotein m RNA levels likewise remained constant despite selenium supplementation. The selenium-dependent change in distribution between the two selenocysteine t RNA isoacceptors also occurred in a cell line-specific manner. The percenta ge of the methylated isoacceptor, mcm(5)Um, changed from 35.5 to 47.2 for D 54, from 38.1 to 47.3 for U251, and from 49.0 to 47.6 for U87. These data r epresent the first time that selenium-dependent changes in selenoprotein mR NA and protein levels, as well as selenocysteine tRNA distribution, were ex amined in human glioma cell lines. BIOCHEM PHARMACOL 60;4:489-497, 2000. (C ) 2000 Elsevier Science Inc.