抑制Wnt信号是生物医学领域关注的焦点问题。反馈抑制是体内抑制Wnt信号的重要方式，而果蝇Nkd即是Wg信号通路的一种反馈抑制因子。尽管Wg信号广泛诱导Nkd的表达，但一般认为Nkd对Wg的拮抗作用仅限于胚胎发育阶段。我们最近在果蝇翅发育模型中阐明了Dpp信号通过Brk-nkd的负调控环路抑制Wg通路的新模型(Yang et al., 2013)，说明nkd在发育后期存在必要作用。本课题将用果蝇遗传学对nkd拮抗Wg信号通路进行深入研究，试图澄清其幼虫发育阶段的作用。同时，我们在果蝇Kc细胞建立了Wg信号通路的定量观察系统，经Wg配体瞬时激活Wg通路对荧光蛋白标记的Nkd和Arm/beta-Catenin进行含量和胞内定位的动态观察，同时利用基于多种Wg靶基因构建的荧光蛋白报告基因实时记录Wg的转录调控活性。通过动力学分析，我们预期可以获得解读Nkd功能的新线索。

The inhibition of Wnt signaling is one of the research focuses in biomedical area. Physiologically, one way to prevent Wnt signaling from overacitivation is achieved by the feedback inhibitory loop. In Drosophila, Nake cuticle (Nkd) is one of such feedback inhibitors thus far been identified for Wingless (Wg, a major fly Wnt) signaling. Previous studies show that the action of Wg signaling appears to perfectly match the sufficient and necessary conditions for Nkd expression in all development stage from egg to adult. However, for a long time, Nkd is thought to only antagnize Wg in certain embryonic development stage and its role in larvae is dispensible. We have recently published a story describing a novel model in which Dpp (a fly BMP) inhibits Wg signaling through brinker-nkd negative circuit in fly wings (Yang et al., 2013), which has certainly shown in one aspect of the essential physiological role of nkd in later devlopmental stage. In this grant we propose to examine the role of nkd on Wg signaling in larval development in more detail aiming to clearify the role of nkd in larval development. In addition to using fly genetics as a major tool, we planed to dig in detail the molecular mechanism by which Nkd influence Wg transcriptional activation based on our newly deviced quantitative Wg activation analytical system. We have constructed Armadillo (Arm, fly beta-catenin) fused to fluoresent protein and Wg reporters from several direct Wg targets. By transient turning on Wg signaling using Wg molecules and manupilation of nkd gene dosage we expect to look into the Nkd nulear localization, Arm stability and the dynamics of Wg transcriptional activation in a more careful way than previously and gain further insights into Nkd function.