Class-Wide Analysis\nof Frizzled-Dishevelled Interactions\nUsing BRET Biosensors Reveals Functional Differences among Receptor\nParalogs

Wingless/Int-1 (WNT) signaling is mediated by WNT binding\nto 10\nFrizzleds (FZD<sub>1–10</sub>), which propagate the signal\ninside the cell by interacting with different transducers, most prominently\nthe phosphoprotein Dishevelled (DVL). Despite recent progress, questions\nabout WNT/FZD selectivity and paralog-dependent differences in the\nFZD/DVL interaction remain unanswered. Here, we present a class-wide\nanalysis of the FZD/DVL interaction using the DEP domain of DVL as\na proxy in bioluminescence resonance energy transfer (BRET) techniques.\nMost FZDs engage in a constitutive high-affinity interaction with\nDEP. Stimulation of unimolecular FZD/DEP BRET sensors with different\nligands revealed that most paralogs are dynamic in the FZD/DEP interface,\nshowing distinct profiles in terms of ligand selectivity and signal\nkinetics. This study underlines mechanistic differences in terms of\nhow allosteric communication between FZDs and their main signal transducer\nDVL occurs. Moreover, the unimolecular sensors represent the first\nreceptor-focused biosensors to surpass the requirements for high-throughput\nscreening, facilitating FZD-targeted drug discovery.