Abstract
Ferritins are protein nanocages that accumulate inside their cavity thousands of oxidized iron atoms bound to oxygen and phosphates. Both characteristic types of eukaryotic ferritin subunits are present in secreted ferritins from insects, but here dimers between Ferritin 1 Heavy Chain Homolog (Fer1HCH) and Ferritin 2 Light Chain Homolog (Fer2LCH) are further stabilized by disulfide-bridge in the 24-subunit complex. We addressed ferritin assembly and iron loading in vivo using novel transgenic strains of Drosophila melanogaster. We concentrated on the intestine, where the ferritin induction process can be controlled experimentally by dietary iron manipulation. We showed that the expression pattern of Fer2LCH-Gal4 lines recapitulated iron-dependent endogenous expression of the ferritin subunits and used these lines to drive expression from UAS-mCherry-Fer2LCH transgenes. We found that the Gal4-mediated induction of mCherry-Fer2LCH subunits was too slow to effectively introduce them into newly formed ferritin complexes. Endogenous Fer2LCH and Fer1HCH assembled and stored excess dietary iron, instead. In contrast, when flies were genetically manipulated to co-express Fer2LCH and mCherry-Fer2LCH simultaneously, both subunits were incorporated with Fer1HCH in iron-loaded ferritin complexes. Our study provides fresh evidence that, in insects, ferritin assembly and iron loading in vivo are tightly regulated.
Highlights
With over one million insect species on earth [1], there can be no simple generalized description of the iron storage strategies they employ [2,3,4,5,6,7,8,9,10,11]
The Ferritin 1 Heavy Chain Homolog (Fer1HCH) and Ferritin 2 Light Chain Homolog (Fer2LCH) subunits have N-terminal signal peptides that direct them to the endoplasmic reticulum [26,27]; Second, Fer1HCH and Fer2LCH are cross-linked to each other by disulfide bonds, giving rise to a highly organized symmetrical arrangement of 12 Fer1HCH and 12 Fer2LCH subunits in the assembled ferritin complex [28]; Third, the Fer1HCH and Fer2LCH genes share common enhancers being chromosomal neighbors and showing post-transcriptional co-regulation to ensure the provision of roughly equal amounts of subunits [16,21,29]
These regulatory relationships are conserved in other insects besides Drosophila melanogaster [30,31]; Fourth, iron loading into ferritin critically depends on transport from the cytosol to the endoplasmic reticulum, a function likely mediated by the zinc regulated and iron regulated transporter 13 (Zip13) [25]; Fifth, the two subunits Fer1HCH and Fer2LCH have been detected in distinct vesicular compartments at the initial stages of the ferritin biosynthetic process, one hour post-feeding on iron-containing media, suggesting that subunit-specific trafficking and post-translational modifications may precede the formation of the ferritin complex [21]
Summary
With over one million insect species on earth [1], there can be no simple generalized description of the iron storage strategies they employ [2,3,4,5,6,7,8,9,10,11]. The Ferritin 1 Heavy Chain Homolog (Fer1HCH) and Ferritin 2 Light Chain Homolog (Fer2LCH) subunits have N-terminal signal peptides that direct them to the endoplasmic reticulum [26,27]; Second, Fer1HCH and Fer2LCH are cross-linked to each other by disulfide bonds, giving rise to a highly organized symmetrical arrangement of 12 Fer1HCH and 12 Fer2LCH subunits in the assembled ferritin complex [28]; Third, the Fer1HCH and Fer2LCH genes share common enhancers (they are transcriptionally co-regulated) being chromosomal neighbors and showing post-transcriptional co-regulation to ensure the provision of roughly equal amounts of subunits [16,21,29] These regulatory relationships are conserved in other insects besides Drosophila melanogaster [30,31]; Fourth, iron loading into ferritin critically depends on transport from the cytosol to the endoplasmic reticulum, a function likely mediated by the zinc regulated and iron regulated transporter 13 (Zip13) [25]; Fifth, the two subunits Fer1HCH and Fer2LCH have been detected in distinct vesicular compartments at the initial stages of the ferritin biosynthetic process, one hour post-feeding on iron-containing media, suggesting that subunit-specific trafficking and post-translational modifications may precede the formation of the ferritin complex [21]. A recent complementary effort in mosquito cells is likely to provide independent information for the ferritin assembly and secretion processes [32]
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