| 1. Background and SignificanceThe flourishing development of Molecular Biology has brought us the best technological platform for investigating insect growth. Insects undergo the characteristic molting and metamorphosis, which needs the hormones20E and juvenile hormone (JH) comprehensive control. Now the studies provided the insulin and FoxO signaling pathway also participated in insect development process. Different signaling pathway interaction, realize the control of insects together. In the signaling pathway, there are insects specific molting and metamorphosis hormones control and eukaryotes conservative insulin and FoxO signaling pathway. Therefore, clarifying these hormones or proteins function is significant on insect growth molecular mechanism.2. Progress and Pending ProblemsThe interaction of hormones20E and JH controls the development of insects.20E is responsible for beginning the molting and the transformation of the larvae to the pupa, JH adjust the physical activation which participates in the molting and metamorphosis coordinated with20E. At present,20E signaling pathway has been detailed researched and how20E signal controls the development is clear; Instead, JH is little studied, we are not committed the receptors of JH, the signaling pathway is also not clear. The research about the interaction between20E signaling pathway and JH is less.There will be some morphologic and tissues changes in the metamorphosis. The most studied are the reconstruction in salivary glands and midguts. Phosphorylation and dephosphorylation, as the most widely exist to regulate the activations, are not fully studied. How to regulate the midgut reconstruction via phosphatase, is the aim in this paper.The development of insects is controlled by insulin and FoxO. Insulin regulates cell division and death, and finally decided the insect growth. Recent research suggests that the hormones20E and insulin influence each other to control insects size. In fruit flies, increases insulin/insulin growth factor signaling not only speed up the larvae growth speed, but also promote the molting hormone synthesis earlier, thus speeding up the metamorphosis happening. The development of insects is regulated by the hormones network, the interaction of hormones control the whole life cycle of insects. FoxO and the insulin regulation are now one of the hot points in scientific research. We found a leptin receptor related gene witch regulates insulin signaling key genes and FoxO and the function of this gene in insects development is another aim in my paper.3. Methods and Acquired ResultsBy using in vivo and in vitro experiments in an epidermal cell line and in larvae of H. armigera, in conjunction with molecular and cellular techniques, we have investigated how PP6participates in the interaction between20E and PCD in H. armigera. We have characterized another protein called leptin receptor overlapping transcripted (LepROT) regulated the development of insects via FoxO.1. PP6could be upregulated by20E and regulated the PCD in midgut. Programmed cell death (PCD) plays an important role in insect midgut remodeling during metamorphosis. Insect midgut PCD is triggered by the steroid hormone20-hydroxyecdysone (20E) and it is mediated by a series of genes. However, the mechanism by which20E triggers midgut PCD is still unclear. Here, we report a protein phosphatase6(PP6) from Helicoverpa armigera playing roles in midgut PCD. PP6was expressed in the midgut during larval growth and it is significantly increased during metamorphosis. The increase was proven to be regulated by20E. The juvenile hormone analog methoprene has no effect on PP6expression. RNA interference analysis suggests that20E upregulated the PP6transcript levels through the ecdysone receptor EcRB1. PP6knockdown by larval feeding or PP6dsRNA injection resulted in the repression of the midgut PCD during the metamorphic stage. The mechanism was demonstrated to be through the suppression of genes such as Broad (Br), E74a, E75b, HR3, E93, rpr, and caspase, which are involved in20E signaling pathway or midgut PCD. These findings suggest that PP6is involved in the20E signal transduction pathway and participates in the PCD in midgut. 2. LepROT regulates the development of insects via FoxO. LepROT is a housekeeping gene, and is not regulated by the20E, Methoprene and insulin. RNAi of LepROT on the larvae could delay the development of the larvae. The expression and the location of FoxO are regulated by LepROT. In the normal cells. FoxO is expressed in the cytoplasm. When the LepROT is downregulated by RNAi, the FoxO is expressed in the nuclei and cytoplasm, but mainly in the cytoplasm. Overexpression of LepROT leads to the inhibition of LepROT. These results suggest that LepROT regulates the development of insects via FoxO.4. Significance of the Acquired ResultsIn this paper, we applied molecular and cellular techniques to investigate the role of proteins that were involved in PCD and metabolisms. There were much more research about the kinases but little study about phosphatases. We firstly elucidated that PP6participated in the regulation of PCD in midgut at metamorphosis. We firstly identified a protein called LepROT and found that this protein could regulate FoxO signaling pathway. Our work provided theoretical basis for elucidating insect development and also provided a reference for investigating the human health and disease-related physiological mechanisms.
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