Proteomics-Based Approach Identifies Altered ER Domain Properties by ALS-Linked VAPB Mutation.

Area of Science:

• Cell Biology
• Neuroscience
• Proteomics

Background:

• Vesicle-associated membrane protein-associated protein B (VAPB) is an ER transmembrane protein crucial for ER-organelle tethering.
• Mutations in VAPB are linked to amyotrophic lateral sclerosis (ALS), causing protein misfolding, aggregation, and ER disorganization.

Purpose of the Study:

• To conduct a proteomic characterization of the ER proteome associated with VAPB and its ALS-linked P56S mutant.
• To investigate the impact of VAPB mutation on ER domain organization and ER-organelle tethering.

Main Methods:

• Optimized immuno-isolation of different ER domains to identify ER sheet- and tubule-specific proteomes.
• Proteomic analysis of VAPB-associated ER proteome in control and P56S mutant conditions.
• Biochemical assays, immunofluorescence, and proximity ligation assays to validate findings.

Main Results:

• VAPB-associated ER proteome exhibited intermediate ER domain properties, with a specific decrease in tubular characteristics upon P56S mutation.
• VAPB mutation led to VAPB delocalization from ER tubules.
• Proteomics indicated reduced incorporation of proteins involved in ER-organelle tethering, confirmed by proximity ligation assays.

Conclusions:

• VAPB mutation alters the proteomic landscape of the ER, specifically affecting its tubular domain properties.
• The study demonstrates that VAPB mutation impairs ER-organelle tethering, potentially through VAPB delocalization.
• These findings provide insights into the molecular mechanisms underlying ER disorganization in VAPB-associated ALS.