Howard Hughes Medical Institute, Waksman Institute and Department of Molecular Biology and Biochemistry, Rutgers The State University of New Jersey, Piscataway, NJ 08854, USA.
Development. 2013 Feb;140(4):831-42. doi: 10.1242/dev.088534. Epub 2013 Jan 14.
The large atypical cadherin Fat is a receptor for both Hippo and planar cell polarity (PCP) pathways. Here we investigate the molecular basis for signal transduction downstream of Fat by creating targeted alterations within a genomic construct that contains the entire fat locus, and by monitoring and manipulating the membrane localization of the Fat pathway component Dachs. We establish that the human Fat homolog FAT4 lacks the ability to transduce Hippo signaling in Drosophila, but can transduce Drosophila PCP signaling. Targeted deletion of conserved motifs identifies a four amino acid C-terminal motif that is essential for aspects of Fat-mediated PCP, and other internal motifs that contribute to Fat-Hippo signaling. Fat-Hippo signaling requires the Drosophila Casein kinase 1ε encoded by discs overgrown (Dco), and we characterize candidate Dco phosphorylation sites in the Fat intracellular domain (ICD), the mutation of which impairs Fat-Hippo signaling. Through characterization of Dachs localization and directed membrane targeting of Dachs, we show that localization of Dachs influences both the Hippo and PCP pathways. Our results identify a conservation of Fat-PCP signaling mechanisms, establish distinct functions for different regions of the Fat ICD, support the correlation of Fat ICD phosphorylation with Fat-Hippo signaling, and confirm the importance of Dachs membrane localization to downstream signaling pathways.
大型非典型钙黏蛋白 Fat 是 Hippo 和平面细胞极性 (PCP) 通路的受体。在这里，我们通过在包含完整 fat 基因座的基因组构建体中创建靶向改变，以及通过监测和操纵 Fat 通路成分 Dachs 的膜定位，来研究 Fat 下游信号转导的分子基础。我们确定人类 Fat 同源物 FAT4 缺乏在果蝇中转导 Hippo 信号的能力，但可以转导果蝇 PCP 信号。靶向缺失保守基序确定了四个氨基酸的 C 末端基序对于 Fat 介导的 PCP 的某些方面是必不可少的，而其他内部基序则有助于 Fat-Hippo 信号转导。Fat-Hippo 信号转导需要果蝇中的 Casein kinase 1ε 编码的 discs overgrown (Dco)，我们在 Fat 细胞内结构域 (ICD) 中鉴定了候选的 Dco 磷酸化位点，突变该位点会损害 Fat-Hippo 信号转导。通过 Dachs 定位的表征和 Dachs 的定向膜靶向，我们表明 Dachs 的定位会影响 Hippo 和 PCP 通路。我们的结果确定了 Fat-PCP 信号转导机制的保守性，确定了 Fat ICD 不同区域的不同功能，支持 Fat ICD 磷酸化与 Fat-Hippo 信号转导的相关性，并证实了 Dachs 膜定位对下游信号通路的重要性。
