Axonal Transport Deficits during Hereditary Spastic Paraplegia

University Of Illinois At Chicago, Chicago IL

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Abstract

DESCRIPTION (provided by applicant): The hereditary spastic paraplegias (HSP) comprise a genetically heterogeneous group of disorders characterized by progressive bilateral weakness and spasticity of the lower limbs. This hallmark clinical symptom of HSP results from progressive dysfunction and degeneration of upper motor neurons in corticospinal tracts and dorsal column fibers. Over fifteen mutations in structurally and functionally unrelated genes have been identified as causative of HSP. Among these, mutations in the SPG4 locus coding for the microtubule-severing protein spastin represent the most common cause of HSP. Despite these major breakthroughs, pathogenic mechanisms underlying HSP pathogenesis remain unknown. Pathological observations from HSP patients and HSP animal models indicate that neurons affected in HSP degenerate following a dying back pattern, which is characterized by early abnormalities in synapses and distal axons and progressive degeneration of axons. Significantly, recent genetic data demonstrated that reductions in fast axonal transport (FAT) result in such pattern of neuronal degeneration. Moreover, loss of function mutations in the SPG10 locus coding for a specific subunit of the molecular motor protein conventional kinesin lead to HSP, suggesting that abnormalities in FAT might indeed represent a critical event in HSP pathogenesis. Our recent studies demonstrated that nanomolar levels of specific mutant spastin isoforms inhibit FAT in an axon-autonomous manner. Consistent with a role of kinases in the regulation of FAT, pharmacological studies presented in this application indicate that this effect of pathogenic spastin was mediated by the activity of casein kinase 2 (CK2). Complementing these observations, active CK2 was found to inhibit FAT. Moreover, CK2 was found to directly phosphorylate and inhibit the functionality of the molecular motor protein conventional kinesin. Based on these and additional findings herein, we propose that activation of axonal CK2 and inhibition of FAT induced by pathogenic spastin represent critical pathogenic events in HSP. Experiments in this application will characterize effects of pathogenic spastin on FAT. Biochemical, immunochemical, pharmacological and cell biological methods will be used to identify specific axonal cargoes and molecular motors associated with pathogenic spastin. Based on our previous work, lentiviral approaches will evaluate isoform-specific effects of pathogenic spastin in vivo. Finally, biochemical, and cell biological approaches will identify and characterize CK2 targets associated with pathogenic spastin including molecular motors and cytoskeletal proteins. These studies will help identifying molecular components and mechanisms mediating the inhibition of FAT induced by pathogenic spastin. Studies proposed here will characterize pathogenic mechanisms underlying the axonal defects characteristic of HSP. The ultimate goal of this project is to identify novel therapeutic targets in HSP that help prevent distal axonopathy and degeneration of motor neurons.