VAC14 oligomerization is essential for the function of the FAB1/PIKfyve-VAC14-FIG4 complex

Area of Science:

• Cell Biology
• Biochemistry
• Structural Biology

Background:

• The PIKfyve-VAC14-FIG4 complex is crucial for synthesizing the signaling lipid PI(3,5)P2.
• VAC14 forms a pentameric scaffold, but the functional significance of its oligomerization was unclear.

Purpose of the Study:

• To elucidate the role of VAC14 oligomerization in the PIKfyve-VAC14-FIG4 complex.
• To investigate the impact of patient-linked mutations on VAC14 function and complex assembly.

Main Methods:

• Atomic-resolution prediction using AlphaFold2 and cryo-EM data.
• Yeast and human cell-based assays to assess protein function, localization, and complex formation.
• Fluorescence-detection size exclusion chromatography (FSEC) to measure VAC14 oligomerization.

Main Results:

• Identified patient mutations within VAC14-VAC14 interfaces, suggesting a role for oligomerization in disease.
• Demonstrated that VAC14 oligomerization is critical for PIKfyve/Fab1 activity and PI(3,5)P2 generation.
• Showed that patient mutations impair complex formation, VAC14 oligomerization, and localization.

Conclusions:

• VAC14 oligomerization is a key regulatory mechanism for PIKfyve/Fab1 activity.
• Understanding these structural and functional roles provides insights into neurodegenerative diseases linked to VAC14 mutations.
• Stabilizing the VAC14 complex may offer a therapeutic strategy for associated diseases.