Ubiquitination of the Dishevelled DIX domain blocks its head-to-tail polymerization

Area of Science:

• Cellular signaling
• Protein ubiquitination
• Molecular biology

Background:

• Dishevelled (Dvl) protein is central to Wnt signaling, relaying signals via its DIX domain.
• DIX domain polymerization forms signalosomes, crucial for Wnt pathway activation.
• Deubiquitinases (DUBs), like CYLD, regulate Dvl activity by removing ubiquitin modifications.

Purpose of the Study:

• To investigate the impact of specific lysine ubiquitination on the Dvl2 DIX domain polymerization.
• To identify DUBs that interact with ubiquitinated Dvl2 DIX domain and characterize their activity.

Main Methods:

• Generation of milligram quantities of pure human Dvl2 DIX domain mono-ubiquitinated at K54 or K58 using genetically encoded orthogonal protection with activated ligation (GOPAL).
• Assay of DIX domain polymerization in solution.
• DUB profiling to identify enzymes cleaving DIX-ubiquitin conjugates.

Main Results:

• Mono-ubiquitination of Dvl2 DIX at K54 inhibits polymerization.
• Mono-ubiquitination at K58 does not prevent DIX domain oligomerization.
• Identification of 28 DUBs that cleave DIX-ubiquitin conjugates, with 14 showing preference or specificity for DIX54-Ub, including Cezanne and CYLD.

Conclusions:

• Ubiquitination site on the Dvl DIX domain dictates its polymerization status.
• Specific DUBs, including CYLD, may promote Dvl signalosome formation by cleaving specific ubiquitin marks, potentially activating Wnt signaling.
• Revises the understanding of DUBs' role in Wnt pathway regulation.