Ferritin RNA interference inhibits the formation of iron granules in the trophocytes of worker honey bees (Apis mellifera)

Abstract

Iron granules containing superparamagnetic magnetite act as magnetoreceptor for magnetoreception in honey bees. Biomineralization of iron granules occurs in the iron deposition vesicles of trophocytes and requires the participation of actin, myosin, ferritin2, and ATP synthase. The mechanism of magnetoreception in honey bees can be explored by suppressing the formation of iron granules. Toward this goal, we injected double-stranded RNA of ferritin2 and ferritin1 into newly emerged worker honey bees to knock down these genes via RNA interference. We confirmed that mRNA and protein production of the ferritins was inhibited, leading to immature iron granules. Downregulating ferritin2 and ferritin1 , moreover, leads to different deposition morphology of 7.5-nm diameter iron particles, indicating that the two genes play different roles in the formation of iron granules in worker honey bees.

Long-term inhibition of ferritin2 synthesis in trophocytes and oenocytes by ferritin2 double-stranded RNA ingestion to investigate the mechanisms of magnetoreception in honey bees (Apis mellifera)

“…Gel electrophoresis via 1.0% agarose gels was performed to verify the expected target. The dsRNA was diluted with nuclease-free water to a final concentration of 5 μg/μl [ 14 , 21 – 23 ].…”

“…Individuals showing hemolymph leakage after microinjection were discarded. Successfully injected bees were housed in a cage (15x10x12 cm) for 1 h before moving into an incubator set to 34°C (NK system, Nippon, Japan) [ 14 ]. Microinjection was carried out three times every 6 days and the number of survival bees was calculated at 1, 7, 13, and 19 days.…”

“…Ferritin is a hollow globular protein containing heavy chains and light chains. Heavy chains called ferritin1 are important for Fe +2 oxidation and have a relationship with the transportation of 7.5-nm diameter iron particles [ 13 , 14 ]. Light chains called ferritin2 assist in core formation and participate in the formation of 7.5-nm diameter iron particles [ 13 , 14 ].…”

“…RNA interference (RNAi)-mediated gene knockdown has been used to knock down vitellogenin , octopamine receptor , DNA methyl-transferase , insulin receptor substrate , tyramine receptor 1 , naked cuticle , and transferrin by double-stranded RNA (dsRNA) injection or ingestion in adult honey bees [ 15 – 22 ]. Recently, we have successfully knocked down ferritin2 and ferritin1 by the one injection of dsRNA into the hemolymph of honey bees [ 14 ]. The mRNA expression and protein synthesis of ferritin2 and the formation of magnetite were inhibited [ 14 ].…”

“…Recently, we have successfully knocked down ferritin2 and ferritin1 by the one injection of dsRNA into the hemolymph of honey bees [ 14 ]. The mRNA expression and protein synthesis of ferritin2 and the formation of magnetite were inhibited [ 14 ]. The one injection of ferritin2 dsRNA shows an inhibitory effect, but its inhibitory effect did not last long enough for behavioral studies.…”