Pathogenic mechanisms of amyotrophic lateral sclerosis-linked VAPB P56S mutation in the degeneration of corticospinal motor neurons.

Aim: The endoplasmic reticulum (ER)-localized vesicle-associated membrane protein-associated protein B (VAPB) is implicated in many cellular processes, such as ER-organelle tethering, calcium homeostasis, and unfolded protein response. The P56S missense mutation in VAPB has been associated with familial forms of motor neuron diseases such as typical amyotrophic lateral sclerosis (ALS), atypical ALS, and spinal muscular atrophy. However, it has not been determined how the VAPB P56S mutation induces the degeneration of corticospinal motor neurons (CSMNs) in ALS. Methods: Using homozygous knock-in (KI) mice expressing P56S VAPB, we investigated the mutation's pathogenic impacts and underlying mechanisms on the survival and function of CSMNs. We performed a wide variety of assays to examine the behavioral, histological, cellular, and molecular abnormalities of KI mice. Results: Compared with wild-type controls, KI mice showed the downregulated protein level of mutant VAPB, proteinase K-resistant cytoplasmic inclusions of mutant VAPB in CSMNs, abnormal hyperactivity, impaired motor coordination, neuronal loss of CSMNs, and axonal degeneration of pyramidal and corticospinal tracts. Mechanistic studies revealed that the VAPB P56S mutation rendered the mutant protein destabilized and inclusion-prone in cortical neurons, and the proteasomal degradation played a critical role in modulating mutant VAPB's protein level and inclusion formation. In addition, the VAPB P56S mutation disrupted ER-mitochondria contacts, impaired VAPB-PTPIP51 interaction and IP3R-VDAC interaction, elevated cytosolic Ca2+, activated CaMKII, and increased CRMP2 phosphorylation. Moreover, the VAPB P56S mutation activated the IRE1-XBP1/p38 mitogen-activated protein kinase (MAPK)/ c-Jun N-terminal kinase (JNK) pathway, increased tau hyperphosphorylation, and upregulated p53 expression and phosphorylation. Conclusion: These findings demonstrate the progressive degeneration of CSMNs induced by VAPB P56S mutation and indicate the involvement of the Ca2+-CaMKII-CRMP2 and IRE1-p38 MAPK/JNK-tau/p53 pathways in the pathogenic process.