Cross-linker design determines microtubule network organization by opposing motors

Abstract

During cell division, cross-linking motors determine the architecture of the spindle, a dynamic microtubule network that segregates the chromosomes in eukaryotes. It is unclear how motors with opposite directionality coordinate to drive both contractile and extensile behaviors in the spindle. Particularly, the impact of different cross-linker designs on network self-organization is not understood, limiting our understanding of self-organizing structures in cells but also our ability to engineer new active materials. Here, we use experiment and theory to examine active microtubule networks driven by mixtures of motors with opposite directionality and different cross-linker design. We find that although the kinesin-14 HSET causes network contraction when dominant, it can also assist the opposing kinesin-5 KIF11 to generate extensile networks. This bifunctionality results from HSET’s asymmetric design, distinct from symmetric KIF11. These findings expand the set of rules underlying patterning of active microtubule assemblies and allow a better understanding of motor cooperation in the spindle.

Effects of microtubule length and crowding on active microtubule network organization

“…Motors remained bound at microtubule ends for an average of 5 s, allowing them to form asters within 35 min in the absence of an attractive depletion force (Fig. 1C, D), as observed previously in experiments and simulations (Roostalu et al 2018, Henkin et al 2022). Whereas microtubules contract locally into small disconnected asters at a lower microtubule density (Fig.…”

“…We simulated active networks consisting of microtubules and microtubule-crosslinking motors using Cytosim. Microtubules and motors were modelled essentially as described earlier (Nedelec 2007, Henkin et al 2022) (see Fig. 1A and Methods).…”

“…Next, we studied the effect of the microtubule length on active microtubule organization. This was done in the absence of a short-range attractive force between microtubules to mimic in vitro experiments in which the transition between contractile and nematic was studied by varying the tubulin concentration in the absence of crowding agents (Henkin et al 2022). In our simulations, we systematically varied the maximum length of the microtubules and the microtubule density, defined as the total length of all microtubules per area (Fig.…”

“…The simulation space and time can however be reduced significantly using shorter microtubules and a higher end-unbinding rate to compensate for their stronger tendency to form contractile networks, while reproducing the same system tendencies and making experimentally testable predictions. This then allows one to simulate the experimentally observed network transitions with shorter microtubules (Henkin et al 2022).…”

“…Biochemical reconstitutions with purified proteins in vitro have made important contributions to our understanding of active network organization and dynamics (Needleman and Dogic 2017, Koenderink and Paluch 2018, Dogterom and Surrey 2013, Alfaro-Aco and Petry 2015). Networks formed from microtubules and motors can display contractile (Nedelec et al 1997, Foster et al 2015, Hentrich and Surrey 2010, Surrey et al 2001, Torisawa et al 2016) or nematic, extensile behavior (Sanchez et al 2012, Lemma et al 2022, Roostalu et al 2018, Henkin et al 2022). Contraction and expansion are fundamentally opposite activities and the molecular parameters that determine these collective behaviors remain incompletely understood.…”