Deep axonal proteomics of human iPSC-derived neurons by microfluidic separation and DIA-MS

Authors

Sauter, C. M., Sandy, Z., Korneck, M., Albrecht, V., Kraft, M., Sivasubramanian, R., Kuttichova, B., Sterneckert, J., Schoels, L., Davies, A., Hauser, S.

Abstract

One of the defining features of neurons is their compartmentalisation into soma, dendrites and axon. Axons are highly specialised for the transmission of signals over long distances but also exhibit unique vulnerabilities in the context of neurodegeneration, implying a specialised axonal proteome. However, in-depth proteomic characterisation of axons has been limited by the difficulty of isolating pure axonal material in sufficient quantities for conventional mass spectrometry analysis. Here, we combine microfluidic-based axon-soma separation with data-independent acquisition mass spectrometry (DIA-MS) on an Orbitrap Astral mass spectrometer for deep profiling of compartment-resolved proteomes of human induced pluripotent stem cell (iPSC)-derived cortical neurons. We quantify over 9,000 proteins in the somatodendritic compartment and ~6,000 proteins in the axonal compartment, to our knowledge, representing the deepest human axonal proteome reported to date. Differential abundance analysis identifies 1,250 axon-enriched proteins with strong enrichment of axon-related pathways including vesicle-mediated transport, synaptic vesicle dynamics, and cytoskeletal organisation. Extending this workflow to iPSC-derived lower motor neurons, we reveal a core set of 417 axon-enriched proteins shared between cortical and lower motor neurons, alongside subtype-specific axonal signatures whose functional differences become evident only through compartment-restricted analysis. We show broad coverage of genes for the neurodegenerative diseases hereditary spastic paraplegia and amyotrophic lateral sclerosis, establishing a quantitative reference for interpreting how disease-linked mutations may differentially affect axonal versus somatodendritic proteostasis. This workflow and resource are readily applicable to other neuronal subtypes and disease models, paving the way for studying axonal proteome dynamics in health and disease.

Preprint server: bioRxiv
The authors list and abstract were imported from bioRxiv on 20 Jul 2026.

Stats

1-terrible, 9-excellent. How would you rate this preprint? Sign in in to submit your rating.

Post a comment

You need to be signed in to post comments. You can sign in here.

Comments

There are no comments yet.