Iron loaded ferritin secretion and inhibition by CI-976 in Aedes aegypti larval cells

Abstract

Ferritin is a multimer of 24 subunits of heavy and light chains. In mammals, iron taken into cells is stored in ferritin or incorporated into iron-containing proteins. Very little ferritin is found circulating in mammalian serum; most is retained in the cytoplasm. Female mosquitoes, such as Aedes aegypti (yellow fever mosquito, Diptera), require a blood meal for oogenesis. Mosquitoes receive a potentially toxic level of iron in the blood meal which must be processed and stored. We demonstrate by 59Fe pulse-chase experiments that cultured A. aegypti larval CCL-125 cells take up iron from culture media and store it in ferritin found mainly in the membrane fraction and secrete iron-loaded ferritin. We observe that in these larval cells ferritin co-localizes with ceramide-containing membranes in the absence of iron. With iron treatment, ferritin is found associated with ceramide-containing membranes as well as in cytoplasmic non-ceramide vesicles. Treatment of CCL-125 cells with iron and CI-976, an inhibitor of lysophospholipid acyl transferases, disrupts ferritin secretion with a concomitant decrease in cell viability. Interfering with ferritin secretion may limit the ability of mosquitoes to adjust to the high iron load of the blood meal and decrease iron delivery to the ovaries reducing egg numbers.

Ferritin is the key to dietary iron absorption and tissue iron detoxification inDrosophila melanogaster

“…Indeed, in mosquitoes, it was reported that radiolabeled dietary iron was found in secreted ferritin in hemolymph (20). Furthermore, we showed that Drosophila ferritin is essential for tissue iron detoxification by exporting excess iron, which is consistent with previous results of cell studies in mosquitoes (34, 35). Considering that different forms of ferritins exist in mammals and insects, and knockdown of ferritin dramatically impairs the survival of flies, ferritin may turn out to be a promising candidate for the control of insect‐transmitted diseases, such as malaria.…”

Insect transferrins: Multifunctional proteins

“…Mammalian 59 Fe-labeled transferrin fed to A. aegypti in a blood meal was absorbed, and the iron recovered and transported to the tissues by secreted ferritin [32]. These findings were consistent with in vitro studies in A. aegypti larval epithelial CCL-125 cells where a pulse-chase experiment with 59 Fe-citrate increased 59 Fe-ferritin secretion into culture medium, while secreted 59 Fe-Tsf was not detected [32,105]. In S. peregrina, radio-labeled iron attached to SpTsf1 was injected into females and the iron was delivered to the eggs and stored in ferritin [45]; these studies also showed uptake of radiolabeled holo-Tsf1 into eggs.…”

The effect of bacterial challenge on ferritin regulation in the yellow fever mosquito, Aedes aegypti

“…We also confirmed that IRP expression is not sensitive to these treatments by a separate set of experiments (Fig. S2A–B; Geiser et al ., ). However, IRP/IRE binding activity is a sensitive measure of active IRP protein in tissues.…”

“…Cell media samples were concentrated fivefold by centrifuging samples in Centricon® Ultracel YM‐30 centrifugal filter devices (Millipore, Billerica, MA, USA) at 5 000 × g for ∼20 min, 4°C. Cell extracts were prepared as previously described with some modification (Geiser et al ., ). Briefly, cell pellets suspended in hypotonic buffer were frozen at −80°C, thawed on ice and homogenized, three times.…”

“…In insects, hemolymph ferritin is found in mg/L concentration (Yoshiga et al ., ; Ong et al ., ). Current evidence supports that secreted ferritin serves as the primary iron transport protein in insects (Nichol et al ., ; Zhou et al ., ; Geiser et al ., ). Zhou et al .…”