Exploring the Role of Nuclear Insulin Signalling in Drosophila Neural Stem Cell Reactivation

本博士项目主要研究果蝇神经干细胞（NSCs）中是否存在一种新的核胰岛素信号通路，并探讨其是否在NSC再激活中起作用。果蝇NSCs在发育过程中的两波神经发生过程中产生中枢神经系统的神经元和胶质细胞。最初，胚胎NSCs分裂产生幼虫神经系统。然后，在胚胎发生结束时，它们进入有丝分裂休眠或静止的可逆阶段。当幼虫进食时，作为对膳食氨基酸的反应，NSCs重新激活，恢复增殖以产生成虫的神经系统。胰岛素信号是NCS再激活的必要条件和充分条件（Chell & Brand, 2010）。胰岛素信号通路在协调生长、代谢和增殖中起着核心作用，毫不奇怪，它在物种间高度保守。在人HepG2肝癌细胞和小鼠原代肝细胞中，胰岛素受体（InR）进入细胞核并与启动子结合，介导基因表达（Hancock et al., 2019）。这揭示了一个范式转移模型，其中InR与典型的膜局部信号转导平行，通过与染色质相关直接影响基因表达。鉴于胰岛素信号在NSC再激活中的作用，我们正在果蝇中探索这种模型。我在研究中使用的核心技术是果蝇遗传学；免疫组织化学和共聚焦显微镜；Targeted DamID （Southall et al., 2013）染色质分析；测序数据的生物信息学分析；以及使用CRISPR/Cas9系统进行基因组编辑。

This PhD project is focused on investigating whether a novel nuclear insulin signalling pathway is present in Drosophila neural stem cells (NSCs), and exploring whether it has a role in NSC reactivation. Drosophila NSCs generate the neurons and glia of the central nervous system during two waves of neurogenesis during development. Initially, embryonic NSCs divide to generate the larval nervous system. They then enter a reversible phase of mitotic dormancy, or quiescence, at the end of embryogenesis. In response to dietary amino acids when the larva feeds, the NSCs reactivate, resuming proliferation to generate the adult nervous system. Insulin signalling is both necessary and sufficient for NCS reactivation (Chell & Brand, 2010). The insulin signalling pathway has a central role in orchestrating growth, metabolism, and proliferation and, unsurprisingly, it is highly conserved across species. In human HepG2 hepatocellular carcinoma cells, and mouse primary hepatocytes, the insulin receptor (InR) was shown to enter the nucleus and associate with promoters, mediating gene expression (Hancock et al., 2019). This reveals a paradigm-shifting model in which, in parallel with canonical membrane-localised signal transduction, the InR directly effects gene expression by associating with chromatin. Given the role of insulin signalling in NSC reactivation, we are exploring this model in Drosophila. The core techniques I use in my research are Drosophila genetics; immunohistochemistry and confocal microscopy; Targeted DamID (Southall et al., 2013) chromatin profiling; bioinformatic analysis of sequencing data; and using the CRISPR/Cas9 system to generate genomic edits.