Overexpression of wild type and mutated human ferritin H-chain in HeLa cells - In vivo role of ferritin ferroxidase activity

Transfectant HeLa cells were generated that expressed human ferritin H-chai n wild type and an H-chain mutant with inactivated ferroxidase activity und er the control of the tetracycline-responsive promoter (Tet-off), The clone s accumulated exogenous ferritins up to levels 14-16-fold over background, half of which were as H-chain homopolymers. This had no evident effect in t he mutant ferritin clone, whereas it induced an iron-deficient phenotype in the H-ferritin wild type clone, manifested by similar to 5-fold increase o f IRPs activity, similar to 2.5-fold increase of transferrin receptor, simi lar to 1.8-fold increase in iron-transferrin iron uptake, and similar to 50 % reduction of labile iron pool. Overexpression of the H-ferritin, but not of the mutant ferritin, strongly reduced cell growth and increased resistan ce to H2O2 toxicity, effects that were reverted by prolonged incubation in iron-supplemented medium. The results show that in HeLa cells H-ferritin re gulates the metabolic iron pool with a mechanism dependent on the functiona lity of the ferroxidase centers, and this affects, in opposite directions, cellular growth and resistance to oxidative damage. This, and the finding t hat also in vivo H-chain homopolymers are much less efficient than the H/L heteropolymers in taking up iron, indicate that functional activity of H-fe rritin in HeLa cells is that predicted from the in vitro data.