Expression of Deptor Gene and Its Effect on the Early Development of Liver and Gut in Zebrafish

The DEPTOR is the inhibitor of mTORC1 complex, which was compromised with other proteins to regulate gene expression of biological processes. But the function of mTOR signaling in development was poorly understood. In our recently studies, the expression time of deptor was opposite to raptor, which expressed protein was the component of mTORC1 complex. The expression of raptor regulated formation of mTORC1 complex to promote proliferation of progenitor cells of liver and gut. When the expression of raptor decreased, the progenitor cells were differentiated and developed into mature liver and gut. However, the regulation of mTORC1 complex formation during liver and gut development is still unknown. To elucidate this issue, we proposed this one-year project, in which we will carry out experiments as follows: (1) Identification of deptor expression during zebrafish development: using whole mount in situ hybridization, we will identify and compare the expression pattern of deptor and raptor during zebrafish development. Furthermore, we will analysis the expression profiles of deptor by qPCR experiment. (2) Overexpression of deptor: microinjection of deptor mRNA into one-cell stage of zebrafish embryo, we will characterize the expression of deptor and raptor by whole mount in situ hybridization. Furthermore, we will analysis the development of liver and gut using liver-specific GFP transgenic line, Tg(LFABP:GFP), gut-specific DsRed transgenic line, Tg(IFABP:DsRed) and immunohistochemistry staining. (3) Morpholino knockdown of deptor: microinjection of deptor morpholino into one-cell stage of zebrafish embryo, we will characterize the expression of deptor and raptor by whole mount in situ hybridization. Furthermore, we will analysis the development of liver and gut using Tg(LFABP:GFP), Tg(IFABP:DsRed) and immunohistochemistry staining. (4) Morpholino knockdown of deptor and treated with rapamycin: microinjection of deptor morpholino into one-cell stage of zebrafish embryo and treated with rapamycin at 3 dpf, we will analysis the development of liver and gut using Tg(LFABP:GFP), Tg(IFABP:DsRed) and immunohistochemistry staining. These results will open a window to expand our knowledge to comprehensive the biological function of deptor during liver and gut development. The usage of rapamycin during pregnancy will also be concerned.