Two bacterial genera, Xenorhabdus and Photorhabdus, are mutually symbiotic to the entomopathogenic nematodes, Steinernema and Heterorhabditis, respectively. Success parasitism of the nematode-bacterial complex depends on the host immunosuppression by the bacteria via their secondary metabolites. Lrp (Leucine-responsive regulatory protein) is a global transcriptional factor of the bacteria and play a crucial role in the parasitism. However, its regulatory targets to suppress the insect immunity were not clearly determined. This study investigated the regulatory target genes and subsequent secondary metabolites by Lrp in Xenorhabdus hominickii. Lrp expression occurred at the early infection stage in a target insect, Spodoptera exigua. Among eight non-ribosomal peptide synthetase (NRPS1-NRPS8) genes, six gene (NRPS3-NRPS8) expressions were positively correlated with Lrp expression in the infected larvae of S. exigua. Exchange of the Lrp promoter with an inducible promoter altered the production of the secondary metabolites along with alteration of the NRPS expression levels. The immunosuppressive activities of X. hominickii depended on the Lrp expression level. The metabolites produced by Lrp expression possessed the eicosanoid-biosynthesis inhibitors and hemolytic factors. A cyclic dipeptide (= cPF) was produced under Lrp control and identified to inhibit phospholipase A2 activity of S. exigua in a competitive inhibitory manner. These results suggest that Lrp is a global transcriptional factor of X. hominickii and plays crucial role in insect immunosuppression by modulating NRPS expressions.

척추동물과 유사하게 곤충도 인지질분해효소(phospholipase A2)의 촉매 작용으로 다양한 아이코사노이드를 합성한다. 그러나 일련의 아 이코사노이드 생합성과정은 척추동물과 차이를 보이는데, 이는 곤충의 인지질에는 전구물질인 아라키도닉산의 함량이 낮기 때문이다. 대신에 비 교적 풍부하게 존재하는 다가불포화지방산인 리놀레익산을 기반으로 사슬 연장 및 불포화반응으로 아라키도닉산을 합성하여 척추동물과 같이 아이코사노이드 전구물질로 이용하는 것 같다. 이렇게 해서 형성된 아라키도닉산은 다시 척추동물의 cyclooxygenase와 유사한 peroxynectin이 PGH2 형태의 프로스타글란딘(prostaglandin: PG) 전구물질을 형성하게 된다. 이후 여러 이성체 효소들의 특이적 반응에 의해 PGA2, PGD2, PGE2, PGI2, TXB2의 다양한 PG가 생성된다. 반면에 또 다른 형태의 아이코사노이드인 에폭시아이코사트리에노익산(epoxyeicosatrienoic acid: EET)은 척추동물과 유사한 단일산화효소의 산화반응으로 아라키도닉산을 전구물질로 5,6-EET, 8,9-EET, 11,12-EET, 14,15-EET를 형성하게 된다. 그러나 세 번째 아이코사노이드 부류인 류코트리엔(leukotriene)의 경우 곤충 체내 존재는 확인되었지만 생합성 과정은 아직 밝 혀지지 않았다. 이들 아이코사노이드가 곤충의 대사, 배설, 면역 및 생식에 관여하는 생리작용을 중개한다. 따라서 아이코사노이드 생합성 과정을 교란하는 물질 탐색은 새로운 살충제 개발 전략이 된다. 본 종설은 이 가운데 PG의 곤충 면역 중개 기작을 소개한다.

곤충면역학은 주로 위생해충인 모기와 말라리아 (또는 Wolbachia)를 중심으로 많은 연구가 이뤄졌으며, 농업해충분 야에서는 곤충병원선충, 기생봉, Polydnavirus, 백강균, Bt 등에 대한 기주곤충과의 상호작용 연구가 기주면역저하와 관련한 생물적방제 인자에 대한 대안으로써 진행되고 있다. 이 가운데, 세균이 생성하는 Bt와 같은 내독소단백질은 기주곤충의 중장에 존재하는 수용체(cadherin, aminopeptidase N, alkaline phosphatase)와의 상호작용으로 기주에 대한 살충효과가 나타는 것을 설명하고 있으며, 최근에는 막관통단백질인 ATP-binding cassette transporter (ABC transporter)는 해독작용과 더불어 Bt와 수용체간의 상호작용에 관여하고 있음이 조명되고 있다. 따라서 곤충의 ABC transporter에 대한 생리적 기능 구명은 기주곤충과 병원체 상호작용 연구의 새로운 장으로 조명될 수 있을 것이다.

Insect immunity is innate and consists of cellular and humoral immune responses. Cellular immune response usually requires hemocyte-spreading behavior, which is accompanied by cytoskeletal rearrangement. A glycolytic enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), catalyzes an oxidation reaction of glyceraldehyde-3-phosphate to 1,3-biphosphoglycerate in the cytosol. Another function of GAPDH in mammalian cell is to bind C-terminal α-tubulin to facilitate cytoskeletal arrangement. An immunoprecipitation (IP) of viral protein, CpBV-CrV1, against hemocyte protein lysate revealed that CpBV-CrV1 binds to GAPDH, identified by MALDI-TOF analysis. RNA interference (RNAi) of GAPDH significantly suppressed cellular immune response, but neither RNAi of hexokinase nor aldolase suppressed the cellular immune response. A common molecular motif of CpBV-CrV1 and a-tubulin at C-terminal region supported the IP analysis. To test the role of α-tubulin motif in CpBV-CrV1, point mutations of CpBV-CrV1 were applied and resulted in loss of the biological activity of CpBV-CrV1. Furthermore, an immunofluorescence assay indicates CpBV-CrV1 colocalized with a-tubulin in hemocytes collected from Plutella xylostella parasitized by Cotesia plutellae possessing C. plutellae bracovirus (CpBV). This result suggests that GAPDH plays a critical role in hemocyte-spreading behavior during immune challenge, and it is a molecular target of the pathogenic virus.

Insects have a highly efficient innate immunity consisting of cellular and humoral responses. Upon microbial pathogen infection, pattern recognition receptors specifically recognize pathogen types and trigger the programmed immune responses via immune mediators, such as cytokine, biogenic monoamines, and eicosanoids. In addition to these canonical immune mediators, two insect developmental hormones of juvenile hormone (JH) and 20-hydroxyecdysone (20E) also modulate immune responses. JH suppresses a cellular immune response, while 20E antagonizes the JH action. In Tribolium castaneum, JH mediates cellular immune responses via non-nuclear receptor(s). When Met (the JH nuclear receptor) expression was silenced by its specific double-stranded RNA, T. castaneum underwent precocious metamorphosis. Under these RNA interference (RNAi) conditions, JH still mediates the cellular immune responses. The addition of calphostin C (a PKC inhibitor) or ouabain (a Na + -K + ATPase inhibitor) significantly suppressed the JH mediation on the cellular immune responses, suggesting the presence of a novel JH receptor(s) at nonnuclear regions. Eicosanoids are a group of oxygenated C20 polyunsaturated fatty acids. They mediate various insect physiological processes including reproduction, excretion, and immunity. Arachidonic acid (AA) is a main precursor of eicosanoid biosynthesis. Phospholipase A2 (PLA2) catalyzes AA release from phospholipids. Subsequent oxygenases of cyclooxygenase (COX) or lipoxygenase (LOX) transform AA to prostaglandins (PGs) or leukotrienes (LTs). In a model insect, Spodoptera exigua, both PGs and LTs mediate both cellular and humoral immune responses. In addition to the physiological significance of the secretory PLA2 (sPLA2), a recent genome analysis identified a novel cellular PLA2 (cPLA2) in S. exigua, which mediates the immune responses. Especially, eicosanoids mediate prophenoloxidase (PPO) activation by stimulating release of PPO from oenocytoid hemocytes. A G protein-coupled receptor is identified in oenocytoids and highly specific to PGE2. Subsequent calcium and PKC pathways mediate PG signal to activate a specific ion channel, Na + -K + -Cl - -cotransporter (NKCC), which generates an osmotic shock to induce cell lysis of oenocytoids to release internal PPO. The released PPO would be activated by a cascade of serine proteases and involved in various immune responses. Eicosanoids also mediate antimicrobial peptide gene expression in response to the bacterial infection. Lack of eicosanid biosynthesis results in a significant immunosuppression. This kind of immunosuppression is exploited by an entomopathogenic bacterium, Xenorhabdus nematophila, to proliferate in the infected larvae without adverse attack to the bacteria by host immune defense. Eight PLA2 inhibitors have been chemically identified and used to develop novel biopesticides. Even though several chemical analyses using various insect systems indicate PG and LT eicosanoids, COX and LOX genes are not identified. Furthermore, little information is available in PG receptors in insects. Thus, signal transduction of eicosanoids in insects remains unresolved. Eicosanoid biosynthesis and signal transduction need to be explored to understand the significant roles of eicosanoids in insect physiological processes.

Phospholipase A2 (PLA2) catalyze the committed step for eicosanoid biosynthesis and releases arachidonic acid (AA), which is oxygenated into eicosanoids that mediate immune responses in insects. Thus, any inhibition of PLA2 activity would lead to a significant immuno suppression due to lack of eicosanoids. Among more than 15 families of PLA2s, group Ⅳ cytosolic PLA2 (cPLA2) has been mainly associated with the production of eicosanoids associated with immune responses. However, no cPLA2 has been reported in all invertebrates including insects. AcPLA2 candidate gene (SecPLA2) has been identified from a hemocyte transcriptome of the beet armyworm, Spodoptera exigua. RNA interference of SecPLA2 expression significantly reduced cellular immune responses of hemocytes. When the SecPLA2 was expressed and purified, the recombinant SecPLA2 catalyzed a substrate, phosphoatidyl choline, atsn-2 position. Its catalytic activity was sensitive to pH, temperature, and calciumlevel. Furthermore, there combinant SecPLA2 was specifically sensitive to a cPLA2-specificinhibitor, methyl arachidonyl fluorophosphonate.