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PLP/DM20 expression and turnover in a transgenic mouse model of Pelizaeus-Merzbacher disease
Saadia A Karim1, Jennifer A Barrie, Mailis C McCulloch
1The Beatson Institute for Cancer Research, Garscube Estate, Switchback Road, Bearsden, Glasgow, G61 1BD, Scotland.
Abstract:
The most common cause of Pelizaeus-Merzbacher (PMD) is due to duplication of the PLP1 gene but it is unclear how increased gene dosage affects PLP turnover and causes dysmyelination. We have studied the dynamics of PLP/DM20 in a transgenic mouse model of PMD with increased gene dosage of the proteolipid protein gene (Plp1). The turnover of PLP/DM20 were investigated using an ex-vivo brain slice system and cultured oligodendrocytes. Homozygous mice have reduced PLP translation, markedly enhanced PLP degradation, and markedly reduced incorporation of PLP into myelin. Proteasome inhibition (MG132) prevented the enhanced degradation. Numerous autophagic vesicles are present in homozygous transgenic mice that may influence protein dynamics. Surprisingly, promoting autophagy with rapamycin decreases the degradation of nascent PLP suggesting autophagic vacuoles serve as a cellular storage compartment. We suggest that there are multiple subcellular fates of PLP/DM20 when overexpressed: the vast majority being degraded by the proteasome, a proportion sequestered into autophagic vacuoles, probably fused with endolysosomes, and only a small proportion entering the myelin sheath, where its association with lipid rafts is perturbed. Transgenic oligodendrocytes have fewer membrane sheets and this phenotype is improved with siRNA-mediated knockdown of PLP expression that promotes the formation of MBP+ myelin-like sheets. This finding suggests that RNAi technology is in principle applicable to improve CNS myelination when compromised by PLP/DM20 overexpression.
Insights
Pelizaeus-Merzbacher disease (PMD) involves PLP1 gene duplication, leading to dysmyelination. This study reveals that increased PLP1 gene dosage enhances protein degradation and impairs myelin incorporation, suggesting RNAi therapy potential.
Area of Science:
- Neuroscience
- Molecular Biology
- Genetics
Background:
- Pelizaeus-Merzbacher disease (PMD) is primarily caused by duplication of the proteolipid protein 1 (PLP1) gene.
- The precise mechanisms by which increased PLP1 gene dosage lead to dysmyelination remain incompletely understood.
Purpose of the Study:
- To investigate the dynamics of PLP/DM20 protein turnover in a transgenic mouse model of PMD with elevated PLP1 gene dosage.
- To elucidate the subcellular fates of overexpressed PLP/DM20 and assess the potential of therapeutic interventions.
Main Methods:
- Utilized an ex-vivo brain slice system and cultured oligodendrocytes from transgenic PMD mice.
- Investigated protein turnover using proteasome inhibition (MG132) and autophagy modulation (rapamycin).
- Assessed myelin formation and oligodendrocyte morphology, including the impact of siRNA-mediated knockdown of PLP expression.
Main Results:
- Homozygous transgenic mice exhibited reduced PLP translation, significantly increased PLP degradation, and diminished incorporation into myelin.
- Proteasome inhibition partially rescued PLP degradation, while rapamycin treatment decreased nascent PLP degradation, suggesting autophagic vacuoles act as storage.
- Overexpressed PLP/DM20 undergoes proteasomal degradation, sequestration into autophagic vacuoles, and limited incorporation into myelin with perturbed lipid raft association.
- Transgenic oligodendrocytes showed reduced membrane sheets, an effect improved by PLP knockdown, promoting MBP+ myelin-like sheet formation.
Conclusions:
- Increased PLP1 gene dosage in PMD leads to dysmyelination through enhanced protein degradation and impaired myelin integration.
- Autophagic vacuoles may serve as a compensatory storage mechanism for excess PLP/DM20.
- RNAi technology targeting PLP1 expression shows promise for improving central nervous system myelination in PMD.
