Rad23B Delays Ataxin-3 Liquid-to-solid Phase Transition Through Heterotypic Buffering

Area of Science:

• Neuroscience
• Biochemistry
• Molecular Biology

Background:

• Abnormal polyglutamine (polyQ) expansion in Ataxin-3 causes Machado-Joseph disease, leading to neuronal nuclear inclusions.
• Ataxin-3 aggregation mechanisms, including amyloid formation and liquid-liquid phase separation (LLPS), are not fully understood.
• Ataxin-3's most abundant brain isoform contains three ubiquitin-interacting motifs (UIMs).

Purpose of the Study:

• Investigate the LLPS properties of the three-UIM Ataxin-3 isoform.
• Clarify the molecular mechanisms of Ataxin-3 aggregation via amyloid formation and LLPS.
• Examine the role of Rad23B interaction in Ataxin-3 aggregation and cellular stress responses.

Main Methods:

• Studied LLPS of Ataxin-3 variants (Q25, Q54, C-terminal fragment) in vitro.
• Assessed droplet aging and amyloid fibril formation using biophysical techniques.
• Investigated heterotypic interactions between Ataxin-3 and Rad23B.
• Observed Ataxin-3 behavior in stress granules under arsenite exposure.

Main Results:

• Ataxin-3 Q25 forms aging phase-separated droplets that mature into amyloid-like fibrils.
• Expanded polyQ (Q54) Ataxin-3 forms short-lived droplets with rapid maturation.
• Ataxin-3 C-terminal fragment forms amyloid fibrils with low Thioflavin-T binding.
• Rad23B inhibits Ataxin-3 droplet maturation but not amyloid formation under dilute conditions.
• Ataxin-3 incorporates into liquid-like stress granules during arsenite stress.

Conclusions:

• Ataxin-3 aggregation via misfolding (amyloid pathway) is distinct from its condensation pathway (LLPS).
• Rad23B modulates Ataxin-3 LLPS but not its amyloidogenic potential under dilute conditions.
• Ataxin-3 aggregation dynamics and stress response roles are elucidated by its behavior in stress granules.