Immune priming is an increased immunity after prior exposure to a specific pathogen as a kind of adaptive immunity and occurs in insects. However, its underlying mechanism is elusive in insects. Immune priming was detected in a lepidopteran insect, Spodoptera exigua. Prior infection with a heat-killed pathogenic bacterium, Xenorhabdus hominickii, increased survival upon the second infection of the live bacteria compared to larvae without pre-exposure. Plasma collected from larvae with the prior infection significantly up-regulated cellular and humoral immune responses compared to the similar treatment without prior exposure. However, when the active plasma exhibiting immune priming was heat-treated, it lost the priming activity, suggesting a presence of protein factor(s) in the immune priming. Lipocalin is a lipid carrier protein and is well known in vertebrates for diverse physiological functions including immunity. An apolipoprotein D3 (ApoD3) is known to be a lipocalin functioning in immune priming in a mosquito, Anopheles gambiae. A homologous ApoD3 (Se-ApoD3) was identified in S. exigua. Se-ApoD3 was expressed in all developmental stages and larvae, it was highly expressed in hemocytes. RNA interference (RNAi) of Se-ApoD3 expression was performed by injecting its specific dsRNA. The larvae treated with the RNAi were impaired in cellular and humoral immune responses. Furthermore, the plasma collected from RNAi-treated larvae lost the immune priming even at the prior exposure. These suggest that Se-ApoD3 mediates the immune priming in S. exigua.

The entomopathogenic bacteria Xenorhabdus/Photorhabdus inhibit insect immune responses by inhibiting eicosanoid biosynthesis. Especially, the bacterial secondary metabolites inhibit PLA2 that release eicosanoid biosynthesis precursors. Some organic extracts of the bacteria-cultured broth possessed PLA2 inhibitory activity. This study used butanol to extract the metabolites and showed that the extract possessed potent inhibitory activity against insect immunity. The inhibition was reversed by the addition of eicosanoid biosynthesis precursor, suggesting PLA2 inhibition. Furthermore, sub-fractionation of the butanol extract separated the secondary metabolites and the fractions exhibited differential PLA2 inhibition. The active fractions appeared to contain novel compounds that are not known in PLA2 inhibition of these bacteria.

An immune signal pathway using Toll receptor in response to Gram positive bacterial or fungal challenge induces cellularimmunity and production of antimicrobial peptides (AMPs) in insects. Here we tested an hypothesis that Toll signal downstreamfactors, MyD88 and Pelle, influence eicosanoid signal by activating phospholipase A2 (PLA2) activity in the beet armywormSpodoptera exigua. We knocked-down MyD88 and Pelle expressions using specific dsRNAs and assessed their effect onPLA2 activity in immune-challenged S. exigua larvae. The RNAi-treated larvae failed to induce AMP gene expressionand PLA2 activity under immune challenge. These results shed light on cross-talk between Toll pathway and eicosanoidsignal.

아이코사노이드는 곤충의 다양한 세포성 면역 반응을 중개한다. 본 연구는 면역반응에 따라 혈구세포 밀도 변화에 대한 아이코사노이드의 새로운 중개 기능을 밝히기 위해 수행되었다. 파밤나방(Spodoptera exigua) 5령충은 세균 감염에 따라 2 시간이 지나면 총혈구수의 현격한 증가를 보였다. 이 총혈구수 증가는 주로 부정형혈구와 소구형혈구 밀도의 증가로 해석되었다. 파밤나방 유충에 phospholipase A2 (PLA2) 억제자인 dexamethasone을 처리하면 세균 처리에 의한 총혈구수 변화가 일어나지 않았다. 하지만 dexamethasone을 처리한 유충에 PLA2의 촉매산물인 arachidonic acid를 첨가하면 총혈구수 증가가 회복되었다. 이러한 혈구 밀도 변화에 원인으로서 아이코사노이드 종류를 추적하기 위해 cyclooxygenase (COX)의 억제자인 naproxene을 처리한 결과 총혈구수 증가가 억제되고, lipoxygenase (LOX)의 억제자인 esculetin을 처리하면 총혈구수 증가가 유지되어 COX 산물이 세균 침입에 따른 총혈구수 증가에 관여하는 것으로 나타났다. COX의 생산물인 prostaglandin E2(PGE2)를 세균 없이 단독으로 처리할 때도 총혈구수의 뚜렷한 증가를 나타냈다. 이러한 결과는 파밤나방의 세포성 면역반응 과정에서 총혈구수 증가를 중개하는 아이코사노이드의 새로운 기능을 제시하고 있다.

Insect blood cells (hemocytes) play a key role in defense against parasites and other pathogenic organisms that infect insects. Cellular immune responses exhibited by hemocytes are acute and effective to initially suppress pathogenic processes. Subsequently humoral immune responses executed by antimicrobial peptides completely cleared the pathogens with help of hemocytes. Two immune mediators, plasmatocyte-spreading peptide (PSP) and eicosanoid, are known to mediate cellular immune responses by activating hemocyte behavior. This study was focused on how these two immune mediators work together to express hemocyte spreading behavior. Both PSP and prostaglandins stimulate hemocyte spreading in dose-dependent manners in the beet armyworm, Spodoptera exigua. Interstingly, inhibition of eicosanoid biosynthesis inhibited PSP activity on mediating the hemocyte-spreading behavior. However, the addition of eicosanoid biosynthesis precursor, arachidonic acid, rescued the hemocytespreading activity. Inhibition of PSP or its receptor by each RNA interference are now under investigation to test whether PSP triggers eicosanoid signaling. These results suggest that there is a cross-talk between PSP and eicosanoid to express hemocyte-spreading behavior in response to bacterial challenge

An entomopathogenic bacterium, Xenorhabdus nematophila, induces an immunosuppression by inhibiting phospholipase A2 (PLA2), which results in a fatal septicemia. PLA2 is an enzyme responsible for eicosanoid biosynthesis and the pathogenic molecular target of this bacterium. A PLA2 gene of the red flour beetle, Tribolium castaneum, was expressed in Escherichia coli. The recombinant T. castaneum PLA2 (TcPLA2) showed enzyme activity, which was specifically inhibited by bromophenacyl bromide (specific inhibitor to secretory PLA2) and ditheothreitol (reducing agent of disulfide bond). It was sensitive to pH (optimum at pH 7.0), temperature (optimum at 30°C), substrate specificity and calcium dependency. X. nematophila released compound(s) inhibiting TcPLA2during its stationary growth phase. The active compound (s) was resistant to heat treatment and could be extracted separately into both organic and aqueous phases. This PLA2 inhibitory fraction showed significant effect on immunosuppression of T. castaneum. These results suggest there may be several PLA2 inhibitors synthesized by X. nematophila and released into culture broth. The recombinant TcPLA2 was also used to screen potent PLA2 inhibitor compounds, which were designed based on a common chemical structure (a pentenebenzene ring) of two peptide inhibitors, proline-tyrosine (PY) and acetylated phenylalanine-glycine-valine (AcFGV). Alterations were made on amino acid sequence or specific functional groups on the pentenebenzene ring. Among 7 different peptides, AY and FGV showed the most potent effects on TcPLA2activity and also resulted in significant reductions in hemocyte spreading behavior of Plutella xylostella. The potent candidate molecules would be applied to control various insect pests to be developed into novel insecticides.

Acyl-CoA synthetase 4는 생쥐에 있어서 거의 모든 조직에서 발현하며 아라키돈산에 특이적인 효소이다. 아라키돈산은 세포막의 인지질로부터 cPLA2에 의하여 유리되고 cyclooxygenase-1, -2에 의하여 eicosanoid로 변환된다. 이렇게 생산된 prostaglandin과 같은 eicosanoid는 배란, 수정, 임신에 있어서 중요한 기능을 수행하고 있다. 그러나 세포막으로부터 유리된 아라키돈산은 acyl-CoA syntheta