PEX1^G843D remains functional in peroxisome biogenesis but is rapidly degraded by the proteasome

Area of Science:

• Cell Biology
• Molecular Genetics
• Biochemistry

Background:

• Peroxisome biogenesis and maintenance rely on the PEX1/PEX6 AAA-ATPase complex.
• Mutations in PEX1 and PEX6 cause Peroxisome Biogenesis Disorders (PBDs), with HsPEX1 G843D being the most common disease-causing mutation.
• The HsPEX1 G843D mutation impairs peroxisomal matrix protein import.

Purpose of the Study:

• To investigate the molecular mechanisms underlying the HsPEX1 G843D mutation's impact on peroxisome function.
• To explore the role of PEX1/PEX6 assembly and stability in PBD pathogenesis.
• To evaluate protein stabilization as a therapeutic approach for PEX1-related PBDs.

Main Methods:

• In vitro biochemical assays using yeast Pex1 G700D mutant to assess ATPase activity and Pex6 assembly.
• Generation and analysis of human cell lines expressing HsPEX1 G843D.
• Proteasomal degradation assays and overexpression studies.
• Analysis of PEX1's affinity for PEX6.
• Deubiquitinase fusion experiments in mammalian cells.

Main Results:

• The yeast Pex1 G700D mutant showed reduced stability and impaired Pex6 assembly but retained ATPase activity, with only minor in vivo import defects.
• Human PEX1 G843D is rapidly degraded by the proteasome, but overexpression can restore peroxisome import.
• The G843D mutation decreases PEX1's affinity for PEX6, and impaired assembly induces degradation of wild-type PEX1.
• Fusing a deubiquitinase to PEX1 G843D significantly reduced its degradation in mammalian cells.

Conclusions:

• Impaired PEX1/PEX6 assembly, rather than solely ATPase dysfunction, is a key driver of PEX1 G843D-associated PBDs.
• Proteasomal degradation of PEX1 is regulated by its assembly with PEX6.
• Stabilizing PEX1 protein, particularly the G843D mutant, presents a promising therapeutic strategy for PBDs caused by PEX1 hypomorphs.