Mechanisms of recognition of non-self and initiation of defense reactions in insects

该研究项目的最终目的是研究入侵昆虫的细菌、真菌和寄生虫等异物被识别为非我的机制，以及非我的识别机制如何与昆虫防御反应的启动相联系。本研究以家蚕血淋巴和体壁两种昆虫组织为研究对象，发现血淋巴中存在酚氧化酶原级联反应。该级联反应由微量的β-1，3-葡聚糖（β G）或肽聚糖（PG）触发。我们之前已经表明，PG和β G在分别特异性结合肽聚糖识别蛋白（PGRP）和β-1，3-葡聚糖识别蛋白（β GRP）时触发级联反应。尽管推测昆虫血腔中的识别蛋白可能在识别细菌或真菌为非我中起着核心作用，但我们对昆虫血腔中识别蛋白的化学结构和作用方式的了解， 关于我们 f识别蛋白是非常有限的。我们用免疫化学和免疫细胞化学方法研究了血腔中识别蛋白的定位，并提出了血腔中两种识别蛋白在细菌或真菌识别非我过程中的作用的工作假说：proPO级联反应的组分包括两种以上的丝氨酸酶酶原。实验结果表明，酶原的活性形式或在其活化过程中由酶原衍生的片段调节血细胞的吞噬活性和血液动力学活性。为了明确证明这种可能性，我们目前正试图纯化所有的proPO级联组件。我们最近已经成功地纯化了proBAEEase及其活性形式至均一。这种活性酶是否对血细胞的吞噬或血细胞动力学活性有影响还有待研究。昆虫体壁的角质层通常被认为是机体的保护性盔甲和化学惰性（死亡）部分。利用家蚕无菌饲养的幼虫，我们可以证明，当角质层的磨损部分有细菌细胞壁成分（脂多糖或肽聚糖）时，角质层向珠被表皮细胞传递信号，指导它们合成抗菌肽（天蚕素）。抗菌肽被分泌到表皮。磨损角质层上的细菌细胞壁成分的存在被证明会导致脂肪体中天蚕素mRNA的合成，但在这种情况下，在脂肪体和血淋巴中没有检测到天蚕素活性。似乎这种mRNA的合成没有立即翻译的信息表明，表皮磨损与细菌细胞壁成分使整个昆虫身体警惕细菌感染。目前，我们正在研究从带有细菌细胞壁成分的角质层磨损部分发送到表皮细胞和脂肪体的信号的性质。少

The ultimate end of this research project is to study the mechanism by which foreign materials such as bacteria, fungi and parasites invading insects are recognized as non-self and how the recognition mechanism of non-self is linked to the initiation of defense reactions of insects. To reach the end the present research was carried out using two insect tissues, hemolymph and integument of the silkworm, Bombyx mori.The prophenoloxidase cascade (ProPO cascade) is present in hemolymph. The cascade is triggered by minute amounts of beta-1,3-glucan (beta G) or peptidoglycan (PG). We have shown previously that PG and betaG trigger the cascade when they bind specifically to peptidoglycan recognition protein (PGRP) and beta-1,3-glucan recognition protein (beta GRP), respectively. Although it is speculated that the recognition proteins may play a central role in the recognition of bacteria or fungi as non-self in insect hemocoel, our knowledge of the chemical structures and the mode of action o … More f the recognition proteins is very limited. We investigated the localization of the recognition proteins in hemocoel by immunochemical and immunocytochemical methods and put forward a working hypothesis on the role of the two recognition proteins in the process of recognition of bacteria or fungi as non-self in hemocoel.The components of proPO cascade include more than two serine enzyme zymogens. Experimental results so har obtained suggest that active forms of the zymogens or fragments derived from the zymogens during their activation modulate phagocytotic activity and hemokinetic activity of hemocytes. To prove this possibility unambiguously we are currently trying to purify all proPO cascade components. We recently have succeeded in purifying proBAEEase and its active form to homogeneity. It is expected that the active enzyme will be studied whether it has the effect on the phagocytotic or hemokinetic activity of hemocytes.Cuticle of insect integument is often considered to be a protective armor and chemically inert (dead) part of the body. Using axenically raised larvae of silkworm, Bombyx mori we could demonstrate that the cuticle transmit a signal to epidermal cells of integument to direct them to synthesize antibacterial peptide (cecropin) when the cuticle has bacterial cell wall component (lipopolysaccharide or peptidoglycan) on its abraded part. The antibacterial peptide is secreted to cuticle. The presence of bacterial cell wall component on abraded cuticle was shown to cause the synthesis of cecropin mRNA in fat body, but in this case no cecropin activity was detected in fat body and hemolymph. Seemingly this mRNA synthesis without immediate translation of the message indicate that cuticular abrasion with bacterial cell wall component make the the entire insect body alert to bacterial infection. Currently we are investigating the nature of signal sent to epidermal cells and fat body from the abraded part of cuticle with bacterial cell wall component. Less