WNT stimulation induced conformational dynamics in the Frizzled-Dishevelled interaction

Abstract

Frizzleds (FZD1-10) are G protein-coupled receptors containing an extracellular cysteine-rich-domain (CRD) that presents the orthosteric binding site of the 19 mammalian WNTs, the endogenous agonists of FZDs. FZDs signal via a diverse set of effector proteins, of which Dishevelled (DVL1-3) is the most well studied and which acts as a hub for several FZD-mediated signaling pathways. However, the mechanistic details of how FZD-DVL interaction mediate pathway initiation and provide pathway selectivity remain an enigma. Here, we use bioluminescence resonance energy transfer-based assays employing Venus-tagged DVL2 together with NanoLuciferase-tagged FZD5 to investigate the WNT-3A- and WNT-5A-induced dynamics of the FZD-DVL interaction. Our biophysical assessment suggests that the ligand-induced BRET changes over time originate from conformational dynamics in the FZD5-DVL2 complex rather than recruitment dynamics of DVL2 to FZD5. Thus, we suggest that extracellular agonist and intracellular transducers could cooperate with each other through allosteric interaction with FZDs in a ternary complex reminiscent of that of classical GPCRs.One Sentence SummaryAnalysis of the interaction of FZD5 and DVL2 uncovers WNT-induced conformational dynamics of a WNT-FZD5-DVL2 complex.

Structural Basis of Frizzled 7 Activation and Allosteric Regulation

“…Additionally, the identi cation of a conserved, tight layer of residues forming the base of the internal pocket that maintains the same organization upon G protein association suggests that constitutive G protein coupling is not su cient to propagate bidirectional allostery from the bottom to the top of receptors in the absence of an agonist. Nonetheless, agonist stimulation of FZDs triggers a conformational change on the intracellular, transducer binding site of the receptor as shown both for G proteins as well as DVL (39) indicating that WNT-CRD interaction elicits conformational dynamics that are transferred to the transducer interaction site in an allosteric manner. This concept is further supported by diverse assessments of FZD dynamics such as the WNT-induced FZD-CRD dynamics (40) as well as agonist-induced FZD conformational changes (41,42).…”

“…The plasmid encoding DEP-Venus has been described previously (39). Plasmids encoding Gα s -67-RlucII and rGFP-CAAX were kindly provided by Prof. Michel Bouvier (IRIC, Université de Montréal, Canada).…”

Navigating breast cancer complexity: Deciphering the cross talk of Wnt/β catenin and Notch pathways in development and diseases

“…The endocytosis of LRP5/6 and subsequent downregulation of Wnt signaling are caused by DKKs, which can inhibit canonical Wnt signaling by binding to LRP5/6 or creating a complex with Kremen 1 and 2. DKKs are an intriguing downstream target of Wnt/catenin signaling, suggesting a negative feedback mechanism to modulate Wnt signaling 42 . By competing with Frizzled receptors for Wnt ligand binding and suppressing Wnt/-catenin signaling, ROR and RYK function as coreceptors of Wnt ligands in the pathway.…”