School of Bioengineering, Dalian University of Technology, Dalian, Liaoning, China.
State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection and Shenzhen Agricultural Genome Research Institute, Chinese Academy of Agricultural Sciences, Beijing, China.
Insect Sci. 2021 Jun;28(3):692-704. doi: 10.1111/1744-7917.12791. Epub 2020 Aug 13.
The insect group II chitinase (ChtII, also known as Cht10) is a unique chitinase with multiple catalytic and chitin-binding domains. It has been proven genetically to be an essential chitinase for molting. However, ChtII's role in chitin degradation during insect development remains poorly understood. Obtaining this knowledge is the key to fully understanding the chitin degradation system in insects. Here, we investigated the role of OfChtII during the molting of Ostrinia furnacalis, a model lepidopteran pest insect. OfChtII was expressed earlier than OfChtI (OfCht5) and OfChi-h, at both the gene and protein levels during larva-pupa molting as evidenced by quantitative polymerase chain reaction and western blot analyses. A truncated OfChtII, OfChtII-B4C1, was recombinantly expressed in Pichia pastoris cells and purified to homogeneity. The recombinant OfChtII-B4C1 loosened compacted chitin particles and produced holes in the cuticle surface as evidenced by scanning electron microscopy. It synergized with OfChtI and OfChi-h when hydrolyzing insoluble α-chitin. These findings suggested an important role for ChtII during insect molting and also provided a strategy for the coordinated degradation of cuticular chitin during insect molting by ChtII, ChtI and Chi-h.
昆虫组 II 几丁质酶（ChtII，也称为 Cht10）是一种具有多个催化和几丁质结合结构域的独特几丁质酶。已被证明其在蜕皮过程中是一种必需的几丁质酶。然而，ChtII 在昆虫发育过程中几丁质降解中的作用仍知之甚少。获得这些知识是全面了解昆虫几丁质降解系统的关键。在这里，我们研究了 OfChtII 在模式鳞翅目害虫棉铃虫蜕皮过程中的作用。通过定量聚合酶链反应和 Western blot 分析，OfChtII 在幼虫-蛹蜕皮过程中的基因和蛋白质水平上的表达均早于 OfChtI（OfCht5）和 OfChi-h。重组表达的截断型 OfChtII（OfChtII-B4C1）在毕赤酵母细胞中表达并纯化至均一性。扫描电子显微镜观察到重组 OfChtII-B4C1 使紧密堆积的几丁质颗粒松散，并在表皮表面产生孔。它与 OfChtI 和 OfChi-h 协同水解不溶性α-几丁质。这些发现表明 ChtII 在昆虫蜕皮过程中具有重要作用，也为 ChtII、ChtI 和 Chi-h 协调降解昆虫蜕皮过程中的表皮几丁质提供了策略。
