Cherqui A, Alla S, Saguez J, Doury G, Sangwan-Norreel B S, Giordanengo P
Laboratoire de Biologie des Entomophages, UPRES EA 2084, Université de Picardie Jules Verne, 33 rue Saint Leu, 80039 Amiens Cedex, France.
J Insect Physiol. 2003 Dec;49(12):1199-209. doi: 10.1016/j.jinsphys.2003.09.004.
beta-glucuronidase (GUS) is a reporter protein commonly expressed in transgenic plants allowing the visualization of the transformed individuals. In our recent work, we showed that consumption of transformed potato plants expressing this GUS enzyme improves performance of the phloem feeding aphid Myzus persicae. Those results led us to the conclusion that the expression of GUS in potato plants might be responsible for the probiotic effect measured in feeding aphids. In the present paper, artificial diets were used to provide active GUS (10 and 500 microg ml(-1)), inactivated heated GUS (500 microg ml(-1)), glucuronic acid (10, 100 and 500 microg ml(-1)), and bovine serum albumin (500 microg ml(-1)) to M. persicae. Our results reveal that these chemicals provided as food intake might influence the biological parameters of this aphid. Experiments showed a probiotic effect of 500 microg ml(-1) GUS diet, resulting in reduced larval mortality, and increased adult reproduction period and fecundity, which led to an increased population growth potential (r(m)=0.17+/-0.01 versus r(m)=0.12+/-0.03 for aphids fed on control diet). A lower amount of added GUS led to fewer variations, biological parameters being only slightly altered (r(m)=0.14+/-0.03). Statistically similar alterations of the biological parameters were obtained when comparing aphids fed on the diet added with inactivated GUS or the non-structural bovine serum albumin protein (r(m)=0.15+/-0.02 and 0.14+/-0.03, respectively). Feeding assays conducted with glucuronic acid supplemented diets enhanced longevity and nymph production of the adult aphids and reduced larval mortality, resulting in r(m)=0.15+/-0.02 for the highest dose (500 microg ml(-1)). Although 100 microg ml(-1) glucuronate diet did not induce any effect on M. persicae (r(m)=0.12+/-0.03), aphids fed on 10 microg ml(-1) glucuronate diet exhibited unexpected reduced demographic parameters (r(m)=0.10+/-0.03). Immuno-histological analysis showed GUS labeling along the whole digestive epithelium of adults and in various tissues including embryos and bacteriocytes. These results suggest that GUS crosses through the digestive tract. Western blots performed with protein extracts of transformed potato plants expressing the gus gene showed a unique band of molecular weight 76 kDa. On the contrary, in extracts from aphids fed on transgenic potato plants or bred on GUS 500 microg ml(-1) artificial diet, several proteins of lower molecular weight were hybridized, revealing proteolysis of ingested GUS. It is concluded that GUS protein, and more precisely GUS activity, is responsible for the probiotic effects on aphid feeding. The possible pathways of induction of such physiological alterations by GUS are discussed.
β-葡萄糖醛酸酶（GUS）是一种常用于转基因植物中的报告蛋白，可使转化个体可视化。在我们最近的研究中，我们发现食用表达这种GUS酶的转基因马铃薯植株可提高以韧皮部为食的蚜虫——桃蚜的生长性能。这些结果使我们得出结论，马铃薯植株中GUS的表达可能是导致取食蚜虫中所测益生菌效应的原因。在本文中，使用人工饲料向桃蚜提供活性GUS（10和500微克/毫升）、灭活加热的GUS（500微克/毫升）、葡萄糖醛酸（10、100和500微克/毫升）以及牛血清白蛋白（500微克/毫升）。我们的结果表明，作为食物摄入的这些化学物质可能会影响这种蚜虫的生物学参数。实验表明，500微克/毫升GUS饲料具有益生菌效应，可降低幼虫死亡率，延长成虫繁殖期并提高繁殖力，从而增加种群增长潜力（内禀增长率r(m)=0.17±0.01，而取食对照饲料的蚜虫r(m)=0.12±0.03）。添加的GUS量较低时，变化较少，生物学参数仅略有改变（r(m)=0.14±-0.03）。当比较取食添加了灭活GUS或非结构牛血清白蛋白的饲料的蚜虫时，生物学参数的变化在统计学上相似（分别为r(m)=0.15±0.02和0.14±0.03）。用添加葡萄糖醛酸的饲料进行的取食试验提高了成年蚜虫的寿命和若虫产量，并降低了幼虫死亡率，最高剂量（500微克/毫升）时r(m)=0.15±0.02。尽管100微克/毫升葡萄糖醛酸盐饲料对桃蚜没有任何影响（r(m)=0.12±0.03），但取食10微克/毫升葡萄糖醛酸盐饲料的蚜虫表现出意外的种群统计学参数降低（r(m)=0.10±0.03）。免疫组织学分析显示，GUS标记遍布成虫整个消化上皮以及包括胚胎和含菌细胞在内的各种组织。这些结果表明GUS穿过了消化道。对表达gus基因的转基因马铃薯植株的蛋白质提取物进行的蛋白质印迹分析显示出一条分子量为76 kDa的独特条带。相反，在取食转基因马铃薯植株或在500微克/毫升GUS人工饲料上饲养的蚜虫提取物中，几种分子量较低的蛋白质发生了杂交，表明摄入的GUS发生了蛋白水解。得出的结论是，GUS蛋白，更确切地说是GUS活性，是对蚜虫取食产生益生菌效应的原因。文中讨论了GUS诱导这种生理变化的可能途径。
