BPS2026 – Modulation of microtubule lateral contacts by fluorescent taxol broadens the architectural selectivity of doublecortin

Microtubules are dynamic cytoskeleton components involved in many cellular activities including trafficking and cell division. Microtubules are targets for numerous small molecules that stabilize them and perturb their dynamics. Because of the ubiquity of microtubules, these small molecules are experimentally and clinically valuable. However, the extent to which their binding modifies the molecular properties of microtubules—including their protofilament architecture—remains an area of active interest. Doublecortin (DCX) is a microtubule-associated protein that nucleates and stabilizes microtubules and plays an important role in neuronal migration. DCX binds microtubules through two conserved DC domains, NDC and CDC, both of which can bind to microtubules at the inter-dimer interface between protofilaments. This binding site has been hypothesized to support DCX’s nucleation and stabilization activities and also renders it sensitive to the number of protofilaments from which microtubules are built. In vitro, nucleation in the presence of doublecortin typically produces 13-protofilament microtubules, which is the main architecture found in cells. Doublecortin binding has thus been used as a tool to differentiate different microtubule architectures. In this study, we first used TIRF microscopy to show that doublecortin binds to microtubules stabilized by the fluorescent taxol derivative, FChitax-3. We used cryo-electron microscopy to determine the structures of DCX-bound FChitax-3-microtubules. The majority of microtubules in these data are built from 15- and 16-protofilaments. We find that the fluorescent moiety of FChitax-3 inserts at inter-protofilament lateral contacts, thereby explaining its influence on microtubule architecture. DCX binds to FChitax-3-microtubules through NDC and our structure allows visualization of the DC domain adaptation to the smaller inter-protofilament angle found in 15-protofilament microtubules. Together, these data reveal the surprising adaptability of microtubule polymers and their cellular binding partners to modulation by small molecules.