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Nitric oxide biosensor uncovers diminished ferrous iron-dependency of cultured cells adapted to physiological oxygen levels

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Gulsah Sevimli, Matthew J. Smith, Tuba Akgul Caglar, Şükriye Bilir, Melike Secilmis, Hamza Y. Altun, Esra N. Yigit, Fan Yang, Thomas P. Keeley, Roland Malli, Gürkan Öztürk, Giovanni E. Mann, Emrah Eroglu

Original languageEnglish
Article number102319
JournalRedox Biology
PublishedJul 2022

Bibliographical note

Funding Information: We gratefully acknowledge support from Scientific and Technological Research Council of Turkey (Grant 118C242, E.E., T.A.C., H.Y.A., M.S., and G.S.), Integration Projects of Sabancı University ( EPD-2019-1 , EE), Heart Research U.K. (RG2673, G.E.M.), British Heart Foundation ( FS/16/67/32548 , G.E.M.), COST Action CA20121 (G.E.M.), and King's Together Strategic Award (G.E.M.). We thank Professors Helmut Sies and Richard J. Naftalin for their helpful discussions. Publisher Copyright: © 2022

King's Authors


Iron is an essential metal for cellular metabolism and signaling, but it has adverse effects in excess. The physiological consequences of iron deficiency are well established, yet the relationship between iron supplementation and pericellular oxygen levels in cultured cells and their downstream effects on metalloproteins has been less explored. This study exploits the metalloprotein geNOps in cultured HEK293T epithelial and EA.hy926 endothelial cells to test the iron-dependency in cells adapted to standard room air (18 kPa O2) or physiological normoxia (5 kPa O2). We show that cells in culture require iron supplementation to activate the metalloprotein geNOps and demonstrate for the first time that cells adapted to physiological normoxia require significantly lower iron compared to cells adapted to hyperoxia. This study establishes an essential role for recapitulating oxygen levels in vivo and uncovers a previously unrecognized requirement for ferrous iron supplementation under standard cell culture conditions to achieve geNOps functionality.

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