Molecular mechanisms in KIF5A-related neurodegeneration

KIF5A is a neuron-specific kinesin driving anterograde axonal transport. It comprises a motor domain for ATP-dependent microtubule binding, a coiled-coil stalk for dimerization and conformational changes, and a tail domain for cargo/adaptor binding and autoinhibition. Mutations targeting the three KIF5A domains give rise to distinct neurodegenerative diseases, but the processes underpinning such genotype/phenotype heterogeneity are not fully understood, yet. Our aim is to investigate the pathogenetic mechanisms at the basis of KIF5A-associated neurodegeneration by characterizing four disease-associated KIF5A mutants (R17Q, R280C, R864X, N999Vfs39) affecting different domains of the protein. Overexpression in NSC-34 motor neuron-like cells showed an abnormal distribution for R864X and N999Vfs39 KIF5A, that preferentially localized within neurites instead of showing a diffused cytoplasmic distribution, possibly due to an impaired kinesin autoinhibition linked to the loss and the alteration of KIF5A tail domain, respectively. More in detail, while R864X KIF5A appeared diffused within neurites, the N999Vfs39 variant formed p62-positive puncta. Notably, both mutants also displayed limited co-localization with mitochondria, a well-established KIF5A cargo, and partially sequestered WT KIF5A within cell protrusions. Cycloheximide chase in SH-SY5Y neuroblastoma cells evidenced shorter half-life for the R17Q and N999Vfs39 mutants compared to WT KIF5A, hinting at altered protein turnover. Accordingly, proteasomal blockage determined R17Q and N999Vfs39 KIF5A accumulation into detergent-insoluble fractions, indicating that the two mutants are preferentially degraded by the ubiquitin-proteasome system and that proteostasis impairment might promote their deposition into potentially harmful aggregates. Interestingly, most features characterizing the ALS-associated N999Vfs39 mutant were recapitulated to a more severe extent by another frameshift variant, C975Vfs73 KIF5A, linked to a rare neurodevelopmental disorder. Together, our observations indicate that both unique and shared mechanisms are involved in the pathogenesis of KIF5A-related NDs. Acknowledgements: Italian Ministry of Health; European Molecular Biology Organization; Company of Biologists