D-010 | hiPSC-derived motor neuron axons show gaps in the periodical spectrin lattice induced by stress

Cellular and Molecular Neurobiology
Author: Nahir Guadalupe Gazal | Email: nggazal@immf.uncor.edu

Nahir Guadalupe Gazal^1°, María Jose Castellanos‐Montiel^2°, Guillermina Bruno^1°,  Anna Franco^2°, Lale Gursu^2°, Ghazal Haghi^2°, Gilles Maussion^2°,  Thomas Durcan^2°, Nicolas Unsain^1°

^1° Instituto Ferreyra (INIMEC-CONICET-Universidad Nacional de Córdoba), Córdoba, Argentina.
^2° Early Drug Discovery Unit, Montreal Neurologial Institute-Hospital, Canada

The membrane-associated periodic skeleton (MPS) is a periodic protein structure of actin “rings” transverse to the axon, separated every 190 nm by “spacers” of α/β-spectrin tetramers. In mature neurons, the MPS is organized along almost the entire axonal shaft, which correlates with the rather homogeneous distribution and high levels of βII-spectrin in this region. During the maturation of motor neurons derived from human induced pluripotent stem cells (hiPSCs MNs) in culture, an intriguing disruption of the otherwise uniform distribution of βII-spectrin along axons was observed. The βII-spectrin gaps (βIIs gaps) appear as regions devoid of βII-spectrin. Various analyses were performed to evaluate axonal constriction or loss. Interestingly, our results indicate that βII-spectrin is the only protein lacking in these specific regions. Since hiPSCs MNs cannot remain healthy beyond 4 weeks, we hypothesized that cellular stress triggers the formation of βIIs gaps. Remarkably, we observed a significant increase in the occurrence of axons with βIIs gaps in 2-week-old cultures only under stress induced by staurosporine, an inhibitor of protein kinases. STED microscopy was used to examine whether βIIs gaps represent a local loss of MPS. We found that this phenomenon is not reversible and can be pharmacologically modulated. We believe that the study of βIIs gaps will provide valuable insights into the formation and dynamics of the MPS.