Department of Physiology & Pharmacology, Section of Receptor Biology & Signaling, Biomedicum, Karolinska Institutet, S-17165 Stockholm, Sweden.
ACS Sens. 2024 Sep 27;9(9):4626-4636. doi: 10.1021/acssensors.4c00806. Epub 2024 Aug 30.
Wingless/Int-1 (WNT) signaling is mediated by WNT binding to 10 Frizzleds (FZD), which propagate the signal inside the cell by interacting with different transducers, most prominently the phosphoprotein Dishevelled (DVL). Despite recent progress, questions about WNT/FZD selectivity and paralog-dependent differences in the FZD/DVL interaction remain unanswered. Here, we present a class-wide analysis of the FZD/DVL interaction using the DEP domain of DVL as a proxy in bioluminescence resonance energy transfer (BRET) techniques. Most FZDs engage in a constitutive high-affinity interaction with DEP. Stimulation of unimolecular FZD/DEP BRET sensors with different ligands revealed that most paralogs are dynamic in the FZD/DEP interface, showing distinct profiles in terms of ligand selectivity and signal kinetics. This study underlines mechanistic differences in terms of how allosteric communication between FZDs and their main signal transducer DVL occurs. Moreover, the unimolecular sensors represent the first receptor-focused biosensors to surpass the requirements for high-throughput screening, facilitating FZD-targeted drug discovery.
无翅型蛋白/Int-1（WNT）信号由 WNT 与 10 个卷曲蛋白（FZD）结合介导，通过与不同的转导子相互作用，尤其是磷酸蛋白 DVL（Dishevelled），在细胞内传递信号。尽管最近取得了进展，但关于 WNT/FZD 选择性和 FZD/DVL 相互作用中旁系同源物依赖性差异的问题仍未得到解答。在这里，我们使用 DVL 的 DEP 结构域作为生物发光共振能量转移（BRET）技术中的替代物，对 FZD/DVL 相互作用进行了全面的分析。大多数 FZD 与 DEP 形成组成性高亲和力相互作用。用不同配体刺激单分子 FZD/DEP BRET 传感器表明，大多数旁系同源物在 FZD/DEP 界面上是动态的，在配体选择性和信号动力学方面表现出不同的特征。这项研究强调了 FZD 和它们的主要信号转导 DVL 之间的变构通讯在机制上的差异。此外，这些单分子传感器代表了第一个超越高通量筛选要求的受体聚焦生物传感器，为 FZD 靶向药物发现提供了便利。
