The Wnt signalling effector Dishevelled forms dynamic protein assemblies rather than stable associations with...

Area of Science:

• Cell Biology
• Molecular Biology
• Signal Transduction

Background:

• Dishevelled (Dvl) is a key protein in the Wnt signaling pathway, crucial for signal transduction.
• Dvl's cytoplasmic puncta formation is linked to its signaling activity, but their nature is unclear.
• Previous assumptions suggested these puncta might be cytoplasmic vesicles.

Purpose of the Study:

• To investigate the molecular composition and dynamics of Dishevelled (Dvl) protein puncta.
• To determine if Dvl puncta represent cytoplasmic vesicles or protein assemblies.
• To elucidate the mechanism of Wnt signal transduction involving Dvl.

Main Methods:

• Live imaging using Total Internal Reflection Fluorescence (TIRF) microscopy of GFP-tagged Dvl2 (GFP-Dvl2).
• Confocal microscopy and photobleaching experiments.
• Colocalization studies with known vesicle markers and lipid dyes.

Main Results:

• Mammalian Dvl2 puncta did not colocalize with endocytic pathway markers or lipid dyes, refuting a vesicular nature.
• TIRF microscopy revealed Dvl2 puncta exhibit undirected movement, collision, and fusion, indicative of dynamic protein aggregates.
• Photobleaching and confocal microscopy demonstrated a dynamic equilibrium between punctate and diffuse cytoplasmic pools of Dvl2 and Axin.

Conclusions:

• Dvl2 and Axin puncta are dynamic protein assemblies, not cytoplasmic vesicles.
• These protein assemblies are in a state of constant flux with the cytoplasmic pool.
• The dynamic nature of these protein assemblies is critical for Wnt signal transduction.