A nucleotide-triggered molecular switch orchestrating septin polymerization

Grupp, B.; Jaeckel, B.; Guenther, A.; Rehberger, J.; Steimle, S.; Gehrke, J. F.; Sieg, C.; Strittmatter, J.; Wunder, T.; Vomhof, T.; Ruhnke, J.; Schleicher, E.; Gerhardt, S.; Johnsson, N.; Gronemeyer, T.

doi:10.64898/2026.05.07.723466

bioRxiv · 10.64898/2026.05.07.723466

A nucleotide-triggered molecular switch orchestrating septin polymerization

Grupp, B.·Jaeckel, B.·Guenther, A.·Rehberger, J.·Steimle, S.·Gehrke, J. F.·Sieg, C.·Strittmatter, J.·Wunder, T.·Vomhof, T.·Ruhnke, J.·Schleicher, E.·Gerhardt, S.·Johnsson, N.·Gronemeyer, T.

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Abstract

Septins are conserved cytoskeletal GTP-binding proteins that form higher-order structures critical for cytokinesis and polarized growth across opisthokonts. In budding yeast and humans, four homologous septins assemble into hetero-octameric protofilaments through alternating interactions between adjacent G-domains (G-interface) or N- and C-termini (NC-interface), which then polymerize end-to-end into filaments. How nucleotide binding and hydrolysis control filament assembly has remained elusive. We uncover a septin-specific mechanism in which nucleotide binding stabilizes the monomeric G-interface and primes it for protofilament formation. Cryo-EM and DEER spectroscopy reveal nucleotide-induced conformational changes that engage the G-interface and enable an NC-compatible conformation absent in the nucleotide-free state. In vitro binding and reconstitution assays confirm that G-interface formation is strictly nucleotide-dependent and required for subsequent NC-interface assembly. These findings establish unidirectional allosteric signaling from the G-to the NC-interface, revealing the molecular basis for controlled septin protofilament assembly.

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Grupp, B., Jaeckel, B., Guenther, A., Rehberger, J., Steimle, S., Gehrke, J. F., Sieg, C., Strittmatter, J., Wunder, T., Vomhof, T., Ruhnke, J., Schleicher, E., Gerhardt, S., Johnsson, N., Gronemeyer, T.. 2026-05-11. A nucleotide-triggered molecular switch orchestrating septin polymerization. https://doi.org/10.64898/2026.05.07.723466

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