A Comparative Analysis of Bombyx mori (Lepidoptera: Bombycidae) β-fructofuranosidase Homologs Reveals Different Post-Translational Regulations in Glyphodes pyloalis Walker (Lepidoptera: Pyralidae).
A Comparative Analysis of Bombyx mori (Lepidoptera: Bombycidae) β-fructofuranosidase Homologs Reveals Different Post-Translational Regulations in Glyphodes pyloalis Walker (Lepidoptera: Pyralidae).
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家蚕β-呋喃果糖苷酶（β-fructofuranosidase，β-FFase）编码基因BmSuc1调控家蚕幼虫的糖代谢，参与桑树生物碱的抗性。然而，目前还没有关于桑树害虫Glyphodes pyloalis步行者β-FFase同源物的分子或生物化学信息。本论文从G. pyloalis的，并表征了GpSUC1a的表达和定位中肠。中肠β-FFase的活性随时间的延长而降低。BmSUC1和GpSUC1a的活性均得到证实，而重组GpSUC1a的活性较低，而BmSUC1的活性较高。在GpSUC1a中发现了一些推测的N-糖基化位点，但在BmSUC1中没有发现，而BmSUC1中的甲基化程度高于GpSUC1a。这些结果表明，GpSUC1a的翻译后修饰（PTMs）差异支持了β-FFase在这两种桑毛虫中的活性，并且GpSUC1a的激活可能受更复杂的翻译后调节系统的控制。果核幼虫这是首次报道G.首次比较了两种桑食虫B. mori和G. pyloalis。本研究为桑螟的防治提供了新的思路。家蚕是一种纺织业和鳞翅目模式昆虫，是桑树专家。BmSuc1基因是第一个在动物中发现的β-呋喃果糖苷酶（β-FFase）编码基因，β-FFase是家蚕幼虫糖代谢调节的必需蔗糖酶，参与了家蚕对桑树生物碱的抗性。桑螟（Glyphodespyloalis步行者）是一种重要的桑树害虫，给蚕业生产造成严重的经济损失。然而，没有关于G的分子或生化信息。pyloalis β-FFase同系物。本研究对G.获得了pyloalis。基因GpSuc 1a和GpSuc 2c在中肠中表达; GpSuc 2c编码截短的多肽。GpSUC1a在中肠的表达和定位进行了表征。在大肠杆菌和BmN细胞中表达的重组GpSUC1a活性较低，而BmSUC1活性较高。pyloalis和中肠纯化的GpSUC1a均得到证实。提示GpSUC1a的激活可能受G.而BmSUC 1在B中的抗性高于BmSUC 1。森。为了研究翻译后修饰（PTM），使用免疫沉淀法从幼虫中肠纯化GpSUC1a和BmSUC1，并进行LC-MS以进行PTM分析。在GpSUC1a中发现了一些推测的N-糖基化位点，而在BmSUC1中没有发现，而BmSUC1中的甲基化程度高于GpSUC1a，表明这些PTM支持了这两种桑毛虫中β-FFases活性的差异。
The β-fructofuranosidase (β-FFase) encoding gene BmSuc1 regulates the glycometabolism of silkworm larvae, and it participates in the resistance of mulberry alkaloids. However, there is no molecular or biochemical information available about the mulberry pest Glyphodes pyloalis Walker β-FFase homologs. In this paper, we have obtained five β-FFase homologous genes in G. pyloalis and characterized the expression and the localization of GpSUC1a in the midgut. The β-FFase activity in the midgut of G. pyloalis larvae and GpSUC1a were both confirmed, while recombinant GpSUC1a displayed little activity as compared with the higher activity of BmSUC1. Some putative N-glycosylation sites were found in GpSUC1a but none in BmSUC1, while there was more methylation in BmSUC1 than in GpSUC1a. The results indicate that such post-translational modifications (PTMs) are differentially supporting that β-FFase are active in these two mulberry feeding caterpillars, and the activation of GpSUC1a may be controlled by a more complex post-translational regulatory system in G. pyloalis larvae. This is the first report on the characterization of β-FFase genes from G. pyloalis and the first comparison of expression regulation between two mulberry feeding insects B. mori and G. pyloalis. Moreover, this research may provide new ideas for the management of mulberry borers. The silk-spinning and Lepidopteran model insect Bombyx mori (Bombycidae) is a mulberry specialist. The BmSuc1 gene is the first β-fructofuranosidase (β-FFase) encoding gene identified in animals, and β-FFase acts as an essential sucrase for glycometabolism modulation in the silkworm larvae, involved in resistance to mulberry alkaloids. Glyphodes pyloalis Walker (Lepidoptera: Pyralidae) is an important mulberry pest leading to heavy economic loss of sericulture. However, no molecular or biochemical information is available about G. pyloalis β-FFase homologs. In this study, five β-FFase homologous genes in G. pyloalis were obtained. The genes GpSuc1a and GpSuc2c were expressed in the midgut; GpSuc2c encodes a truncated polypeptide. The expression and the localization of GpSUC1a in the midgut was characterized. Whereas recombinant GpSUC1a expressed in both Escherichia coli and BmN cells displayed little activity as compared with higher activity of BmSUC1, β-FFase activity in the larval midgut of G. pyloalis and GpSUC1a purified from the midgut were both confirmed. The data suggested that the activation of GpSUC1a is probably controlled by a more complicated post-translational regulation system in G. pyloalis larvae than that of BmSUC1 in B. mori. To study post-translational modifications (PTMs), GpSUC1a and BmSUC1 were purified from larval midguts using immunoprecipitation and subjected to LC-MS to perform PTMs analysis. Some putative N-glycosylated sites were found in GpSUC1a but none in BmSUC1, while there was more methylation in BmSUC1 than in GpSUC1a, indicating that such PTMs were supporting the differential β-FFases activities in these two mulberry feeding caterpillars.
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作者：
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DOI：
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期刊：
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