| Research background, scientific issues and significanceInsects are the most species, and the largest animal groups exist on the Earth’s land. Unlike vertebrates, their bodies do not have the support of endoskeleton. Their bodies are wrapped in a layer which supports and protects the body as a wall. This hard body walls no ductility, and in the process of the insect growth will hinder the growth of the body, thus the growth and development of insect body must be in the periodic ecdysis process. Insect’s life experience four distinct development period:the egg, larva, pupa and adult, each period has different function and formation. Among which, ecdysis between larva to larva called larva molting(molting), transition from larvae to pupa is called metamorphosis molting(metamorphosis). Insect molting and metamorphosis mainly regulate by ecdysone (20-hydroxyecdysone,20E)and juvenile hormone (JH) together, and it is a complex series of cascade reaction process.20E decided the time interval of molting and induce the death of the old organization; JH antagonize the20E effect and determine the nature of molting.In insect development, the reconstruction of organization accompanies this process. Late in the larvae metamorphosis, larvae’s organization disintegrates, replaced by a new generation of adult tissue. Tissue growth and death under the influence of different internal and external factors, such as hormone and nutrition status, inside is the regulation of cell proliferation and apoptosis. Hippo pathway controls cell growth, reproduction and apoptosis in insects and vertebrates. The prime target of this pathway in growth regulation is transcriptional co-activator Yorkie (Yki), which is an important part of Hippo pathway. The function of Hippo pathway in insect molting and metamorphosis during development and its relationship with20E has’t been reported. In this article, I has studied this issue. Insects are closely related to humans, and have received extensive attention of people since time immemorial. The study of insect molting and metamorphosis is both understanding evolution of a breakthrough, also can further understand the molecular mechanism of insect development, and find new available molecular targets for pest control.Methods and Acquired ResultsIn this paper, I use Helicoverpa armigera as experiment material,employ gene cloning,recombinant protein expression and polyclonal antibody preparation methods to explore Yki protein expression patterns in the Helicoverpa armigera. Throgh20E hormone stimulation in vivo and in vitro, detect the expression changes of Yki when ecdysone implemented. Observation by immunohistochemical finds the distribution of Yki in midgut of cotton bollworm. We get following results:From Helicoverpa armigera, I had got the full length of yki sequence. Currently yki has1320nucleotide as open reading frame, it encodes440amino acids. Sequence alignment results and evolutionary tree analysis showed that the similarity between Helicoverpa armigera yki and Bombyx mori yki is92.5%.Western blot shows that Yki high express in metamorphosis period (72-120h,6th age) of larvae midgut. Hormone stimulation result shows20E can promote Yki protein expression quantity in vivo, but JH can’t. Accordingly, Yki is regulated by20E pathway.Immunocytochemistry showed that Yki was located in both the nucleus and the cytoplasm of normal cells;20E and JH could not promote the translocation of Yki from cytoplasm to the nucleus. Immunohistochemistry results indicate that Yki was distributed in old midgut of bollworm during its metamorphosis.These results suggest that Yki may involve in the bollworm organization reconstruction process.Research conclusions and SignificanceIn this paper, the Helicoverpa armigera acts as the experimental material, revealed Yki protein is highly expressed in larval metamorphosis period, can be raised by20E, and distributed in old midgut, which laid a foundation for further exploration of interaction mechanism of Yki protein with the20E signal pathway. |
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