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Ferric Chelate Reductase Knockdown in Drosophila S2 Cells
Author(s) -
Murphy Laura Grace,
Holst Jessica,
Kane Gregory,
Gorman Maureen J.,
Ragan Emily J.
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.03495
Subject(s) - gene knockdown , rna interference , ferric , reductase , microbiology and biotechnology , drosophila (subgenus) , chemistry , chelation , biology , biochemistry , enzyme , gene , rna , organic chemistry
Although the mechanisms of cellular iron uptake in mammals are largely understood, there is still little known about the mechanism of iron transport in insects. Not only is iron essential, it can also be toxic and therefore needs to be well regulated to maintain balance between necessity and toxicity. In mammals, a duodenal cytochrome b acts as a ferric reductase to allow for uptake of iron into intestinal cells. We identified a cytochrome b561 family member in Drosophila , CG8399, that is expressed in Drosophila S2 cells. CG8399 is similar to ferric chelate reductase‐1 in humans as both contain reeler, domon and cytochrome b561 domains. We are testing the hypothesis that there is reduction of iron by CG8399, which allows the transport of iron into the cells. To test this hypothesis, we used RNA interference (RNAi) to knockdown the CG8399 mRNA. We quantified the RNAi knockdown caused by RNAi using real time PCR and compared iron concentrations between untreated and treated cells using a ferrozine‐based iron content assay. A lower concentration of iron in CG8399 RNAi‐treated cells would support our hypothesis. Support or Funding Information NSF IOS‐1656407