ASCORBIC-ACID ENHANCES IRON-INDUCED FERRITIN TRANSLATION IN HUMAN LEUKEMIA AND HEPATOMA-CELLS

Ascorbate is an important cofactor in many cellular metabolic reaction s and is intimately linked to iron homeostasis. Continuously cultured cells are ascorbate deficient due to the lability of the vitamin in so lution and to the fact that daily supplementation of media with ascorb ate is unusual. We found that ascorbate repletion alone did not alter ferritin synthesis. However, ascorbate-replete human hepatoma cells, H ep3B and HepG2, as well as K562 human leukemia cells achieved a substa ntially higher cellular ferritin content in response to a challenge wi th iron than did their ascorbate-deficient counterparts grown under st andard culture conditions. Most of the elevation in ferritin content w as due to an increase in de novo ferritin synthesis of greater than 50 -fold, as shown by in vivo labeling with [S-35]methionine and immunopr ecipitation. RNA-blot analysis showed only minor changes in steady sta te levels of ferritin mRNA, suggesting that ascorbate enhances iron-in duced ferritin synthesis primarily by post-transcriptional events. Tra nsient gene expression experiments using chloramphenicol acetyltransfe rase reporter gene constructs showed that the ascorbate effect on ferr itin translation is not mediated through the stem-loop near the transl ational start site that transduces ferritin synthesis in response to c ytokines. The data suggest that ascorbate possibly modifies the action of the iron-responsive element on ferritin translation, although more precise structure-function studies are needed to clarify this issue. These data demonstrate a novel role of ascorbate as a signaling molecu le in posttranscriptional gene regulation. The mechanism by which asco rbate modulates cellular iron metabolism is complex and requires addit ional detailed investigation.