Sonicated fibrils of huntingtin exon-1 preferentially seed neurons and produce toxic assemblies
HD is a genetically inherited neurodegenerative disorder caused by expansion of a polyglutamine (polyQ) repeats in the exon-1 of huntingtin protein (HTT). The expanded polyQ enhances the amyloidogenic propensity of HTT exon 1 (HTTex1), which forms a heterogeneous mixture of assemblies with some being neurotoxic. While predominantly intracellular, monomeric and aggregated mutant HTT species are also present in the cerebrospinal fluids of HD patients, however, their biological properties are not well understood. To explore the role of extracellular mutant HTT in aggregation and toxicity, we investigated the possible uptake and amplification of recombinant HTTex1 assemblies in cell culture models. We found seedingcompetent species in the sonicated HTTex1 fibrils, which preferentially entered human neurons and triggered the amplification of neurotoxic assemblies; astrocytes or epithelial cells were not permissive to the HTTex1 seeding. The aggregation of HTTex1 seeds in neurons depleted endogenous HTT protein with non-pathogenic polyQ repeat, activated apoptotic caspase-3 pathway and induced nuclear fragmentation. Using a panel of novel monoclonal antibodies and genetic mutation, we identified epitopes within the N-terminal 17 amino acids and proline-rich domain of HTTex1 mediating neural seeding. Synaptosome preparations from the brains of HD mice also contained similar neurotoxic seeding-competent mutant HTT species. Our findings suggest that amyloidogenic extracellular mutant HTT assemblies may selectively enter neurons, propagate and produce neurotoxic assemblies.
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. This version posted April 18, 2021. We are grateful to Dr. Dr. Alejandro Balazs at the Ragon Institute for providing the IgG2 cDNA backbone and Dr. Alysson Muotri at USCD for the IPSC neuronal progenitors. We are also thankful to Dr. Jenny Morton at the University of Cambridge, Dr. Ignacio Munoz-Sanjuan and Dr. Jonathan Bard at CHDI, and Dr. Jeannie Chen at USC for critical suggestions. Ethics approval: All animal experiments and care complied with federal regulations and were reviewed and approved by the California Institute of Technology Institutional Animal Care and Use Committee (IACUC), protocol#1776-19. Funding was provided by a CHDI award to AK (A-12890), CHDI award (A-12640) to RL and AK, and 1R01NS084345 to RL. Authors' contributions: AK conceived the idea and wrote the manuscript. AC, JMI, NKP, AR, TM performed the experiments. AK, AC, RL and JMI analyzed the data. RL edited the manuscript. All authors have read and approved the paper. Authors declare no competing interest.
Submitted - 2021.04.16.440200v1.full.pdf
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