Tracking Intracellular Labile Iron with a Genetically Encoded Fluorescent Reporter System Based on Protein Stability.

ABSTRACT: Here, we developed IronFist, a genetically encoded fluorescent reporter that enables dynamic tracking of labile ferrous ions (Fe²⁺) in live cells. IronFist is a bicistronic system combining the iron-sensitive hemerythrin-like (Hr) domain from the F-box and leucine-rich repeat protein 5 (FBXL5), fused to the bright fluorescent protein (FP) mNeonGreen, alongside mCherry as a reference FP signal. When labile iron levels are low, Hr-mNeonGreen undergoes ubiquitination and degradation, leading to a low green-to-red fluorescence ratio. Conversely, as cytosolic Fe²⁺ levels rise and Fe²⁺ ions bind to Hr, the green fluorescence is stabilized, increasing the IronFist ratio signal. Using IronFist for end point measurements, we observed that most cells maintain low bas

SUBMITTER: Akyol A 

PROVIDER: S-EPMC12379171 | biostudies-literature | 2025 Aug

REPOSITORIES: biostudies-literature