Dutta Abinash, Dandapat Jagneshwar, Mohanty Nakulananda
P.G. Department of Biotechnology, Utkal University, Bhubaneswar 751004, Odisha, India.
P.G. Department of Biotechnology, Utkal University, Bhubaneswar 751004, Odisha, India.
Comp Biochem Physiol B Biochem Mol Biol. 2019 Jul;233:23-34. doi: 10.1016/j.cbpb.2019.03.010. Epub 2019 Apr 1.
Transferrin is a highly conserved multifunctional protein involved in iron metabolism, oxidative stress and immune response. However, very little is known about its context-dependent functional role in insects. The present study adopts a proteomic approach to identify an abundant hemolymph protein of silkworm (Antheraea mylitta) as transferrin. SDS-PAGE coupled to LC-MS/MS was followed to identify this protein (75.7 kDa) in the advanced larvae of A. mylitta as transferrin (AmTsf). Antibody developed against this protein enabled us to determine its tissue-specific expression and functional relevance during development. Its immunodetection was optimized by western blotting, immunohisto- and cytochemistry in the larval and pupal tissues of this insect. AmTsf exhibited a considerable homology with that of other related insects. It was found to be expressed constitutively in hemolymph, fat body, midgut and silk gland of this insect during development. Abundance of transferrin in the hemolymph of this species and its upregulation in response to oxidant challenges indicated its tissue-specific role in the antioxidant protection. The present work adds to the existing knowledge of multiple roles of transferrin in insects and provides an insight into a relatively unexplored aspect of iron-associated HO metabolism and redox homeostasis in the apparent absence of catalase and glutathione peroxidase. Therefore, A. mylitta can be an attractive acatalasemic model for the study of in vivo redox regulation. In addition, this study provides a novel hypothesis to explore the role of transferrin in limiting oxidative stress during larval rearing of silkworm.
转铁蛋白是一种高度保守的多功能蛋白质，参与铁代谢、氧化应激和免疫反应。然而，关于其在昆虫中依赖于环境的功能作用，人们了解甚少。本研究采用蛋白质组学方法，鉴定出家蚕（柞蚕）一种丰富的血淋巴蛋白为转铁蛋白。随后采用SDS-PAGE与LC-MS/MS联用的方法，在柞蚕大龄幼虫中鉴定出这种75.7 kDa的蛋白质为转铁蛋白（AmTsf）。针对该蛋白制备的抗体使我们能够确定其在发育过程中的组织特异性表达及其功能相关性。通过蛋白质免疫印迹法、免疫组织化学和免疫细胞化学方法，对该昆虫幼虫和蛹组织中该蛋白的免疫检测进行了优化。AmTsf与其他相关昆虫的转铁蛋白具有相当高的同源性。研究发现，在该昆虫发育过程中，转铁蛋白在其血淋巴、脂肪体、中肠和丝腺中组成性表达。该物种血淋巴中转铁蛋白的丰度及其在氧化应激刺激下的上调表明其在抗氧化保护中的组织特异性作用。本研究增加了我们对转铁蛋白在昆虫中多种作用的现有认识，并为在明显缺乏过氧化氢酶和谷胱甘肽过氧化物酶的情况下，铁相关的血红素加氧酶代谢和氧化还原稳态这一相对未被探索的方面提供了见解。因此，柞蚕可能是研究体内氧化还原调节的一个有吸引力的无过氧化氢酶模型。此外，本研究为探索转铁蛋白在限制家蚕幼虫饲养过程中的氧化应激作用提供了一个新的假说。
