Cysteine mutant of mammalian GPx4 rescues cell death induced by disruption of the wild-type selenoenzyme.

Selenoproteins are expressed in many organisms, including bacteria, insects, fish, and mammals. Yet, it has remained obscure why some organisms rely on selenoproteins while others, like yeast and plants, express Cys-containing homologues. This study addressed the possible advantage of selenocysteine (Sec) vs. Cys in the essential selenoprotein glutathione peroxidase 4 (GPx4), using 4-hydroxy-tamoxifen-inducible Cre-excision of loxP-flanked GPx4 alleles in murine cells. Previously, it was shown that GPx4 disruption caused rapid cell death, which was prevented by α-tocopherol. Results presented herein demonstrate that the expression of wild-type (WT) GPx4 and its Sec/Cys (U46C) mutant rescued cell death of GPx4(-/-) cells, whereas the Sec/Ser (U46S) mutant failed. Notably, the specific activity of U46C was decreased by ∼90% and was indistinguishable from U46S-expressing and mock-transfected cells. Hence, the U46C mutant prevented apoptosis despite hardly measurable in vitro activity. Doxycycline-inducible expression revealed that minute amounts of either U46C or WT GPx4 prevented cell death, albeit WT GPx4 was more efficient. Interestingly, at the same expression level, proliferation was promoted in U46C-expressing cells but attenuated in WT-expressing cells. In summary, both catalytic efficiency and the expression level of GPx4 control the balance between cell survival and proliferation.