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1. Lateral and longitudinal compaction of PRC1 overlap zones drives stabilization of interzonal microtubules

   https://doi.org/10.1091/mbc.E23-02-0049  

   do_Rosário, Carline Fermino; Zhang, Ying; Stadnicki, Jennifer; Ross, Jennifer L; Wadsworth, Patricia (September 2023, Molecular Biology of the Cell)

   Wignall, Sarah (Ed.)

   During anaphase, antiparallel–overlapping midzone microtubules elongate and form bundles, contributing to chromosome segregation and the location of contractile ring formation. Midzone microtubules are dynamic in early but not late anaphase; however, the kinetics and mechanisms of stabilization are incompletely understood. Using photoactivation of cells expressing PA-EGFP-α-tubulin we find that immediately after anaphase onset, a single highly dynamic population of midzone microtubules is present; as anaphase progresses, both dynamic and stable populations of midzone microtubules coexist. By mid-cytokinesis, only static, non-dynamic microtubules are detected. The velocity of microtubule sliding also de-creases as anaphase progresses, becoming undetectable by late anaphase. Following depletion of PRC1, midzone microtubules remain highly dynamic in anaphase and fail to form static arrays in telophase despite furrowing. Cells depleted of Kif4a contain elongated PRC1overlap zones and fail to form static arrays in telophase. Cells blocked in cytokinesis form short PRC1 overlap zones that do not coalesce laterally; these cells also fail to form static arrays in telophase. Together, our results demonstrate that dynamic turnover and sliding of midzone microtubules is gradually reduced during anaphase and that the final transition to astatic array in telophase requires both lateral and longitudinal compaction of PRC1 containing overlap zones. 

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2. Focal adhesions are controlled by microtubules through local contractility regulation

   https://doi.org/10.1038/s44318-024-00114-4  

   Aureille, Julien; Prabhu, Srinivas S; Barnett, Sam F; Farrugia, Aaron J; Arnal, Isabelle; Lafanechère, Laurence; Low, Boon Chuan; Kanchanawong, Pakorn; Mogilner, Alex; Bershadsky, Alexander D (July 2024, The EMBO Journal)

   Abstract Microtubules regulate cell polarity and migration via local activation of focal adhesion turnover, but the mechanism of this process is insufficiently understood. Molecular complexes containing KANK family proteins connect microtubules with talin, the major component of focal adhesions. Here, local optogenetic activation of KANK1-mediated microtubule/talin linkage promoted microtubule targeting to an individual focal adhesion and subsequent withdrawal, resulting in focal adhesion centripetal sliding and rapid disassembly. This sliding is preceded by a local increase of traction force due to accumulation of myosin-II and actin in the proximity of the focal adhesion. Knockdown of the Rho activator GEF-H1 prevented development of traction force and abolished sliding and disassembly of focal adhesions upon KANK1 activation. Other players participating in microtubule-driven, KANK-dependent focal adhesion disassembly include kinases ROCK, PAK, and FAK, as well as microtubules/focal adhesion-associated proteins kinesin-1, APC, and αTAT. Based on these data, we develop a mathematical model for a microtubule-driven focal adhesion disruption involving local GEF-H1/RhoA/ROCK-dependent activation of contractility, which is consistent with experimental data. 

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3. Mechanisms of microtubule dynamics and force generation examined with computational modeling and electron cryotomography

   https://doi.org/10.1038/s41467-020-17553-2  

   Gudimchuk, Nikita B.; Ulyanov, Evgeni V.; O’Toole, Eileen; Page, Cynthia L.; Vinogradov, Dmitrii S.; Morgan, Garry; Li, Gabriella; Moore, Jeffrey K.; Szczesna, Ewa; Roll-Mecak, Antonina; et al (July 2020, Nature Communications)

   Abstract Microtubules are dynamic tubulin polymers responsible for many cellular processes, including the capture and segregation of chromosomes during mitosis. In contrast to textbook models of tubulin self-assembly, we have recently demonstrated that microtubules elongate by addition of bent guanosine triphosphate tubulin to the tips of curving protofilaments. Here we explore this mechanism of microtubule growth using Brownian dynamics modeling and electron cryotomography. The previously described flaring shapes of growing microtubule tips are remarkably consistent under various assembly conditions, including different tubulin concentrations, the presence or absence of a polymerization catalyst or tubulin-binding drugs. Simulations indicate that development of substantial forces during microtubule growth and shortening requires a high activation energy barrier in lateral tubulin-tubulin interactions. Modeling offers a mechanism to explain kinetochore coupling to growing microtubule tips under assisting force, and it predicts a load-dependent acceleration of microtubule assembly, providing a role for the flared morphology of growing microtubule ends. 

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4. Active nematic order and dynamic lane formation of microtubules driven by membrane-bound diffusing motors

   https://doi.org/10.1073/pnas.2117107118  

   Memarian, Fereshteh L.; Lopes, Joseph D.; Schwarzendahl, Fabian Jan; Athani, Madhuvanthi Guruprasad; Sarpangala, Niranjan; Gopinathan, Ajay; Beller, Daniel A.; Dasbiswas, Kinjal; Hirst, Linda S. (December 2021, Proceedings of the National Academy of Sciences)

   Dynamic lane formation and long-range active nematic alignment are reported using a geometry in which kinesin motors are directly coupled to a lipid bilayer, allowing for in-plane motor diffusion during microtubule gliding. We use fluorescence microscopy to image protein distributions in and below the dense two-dimensional microtubule layer, revealing evidence of diffusion-enabled kinesin restructuring within the fluid membrane substrate as microtubules collectively glide above. We find that the lipid membrane acts to promote filament–filament alignment within the gliding layer, enhancing the formation of a globally aligned active nematic state. We also report the emergence of an intermediate, locally ordered state in which apolar dynamic lanes of nematically aligned microtubules migrate across the substrate. To understand this emergent behavior, we implement a continuum model obtained from coarse graining a collection of self-propelled rods, with propulsion set by the local motor kinetics. Tuning the microtubule and kinesin concentrations as well as active propulsion in these simulations reveals that increasing motor activity promotes dynamic nematic lane formation. Simulations and experiments show that, following fluid bilayer substrate mediated spatial motor restructuring, the total motor concentration becomes enriched below the microtubule lanes that they drive, with the feedback leading to more dynamic lanes. Our results have implications for membrane-coupled active nematics in vivo as well as for engineering dynamic and reconfigurable materials where the structural elements and power sources can dynamically colocalize, enabling efficient mechanical work. 

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5. Microtubules form by progressively faster tubulin accretion, not by nucleation–elongation

   https://doi.org/10.1083/jcb.202012079  

   Rice, Luke M.; Moritz, Michelle; Agard, David A. (March 2021, Journal of Cell Biology)

   Microtubules are dynamic polymers that play fundamental roles in all eukaryotes. Despite their importance, how new microtubules form is poorly understood. Textbooks have focused on variations of a nucleation–elongation mechanism in which monomers rapidly equilibrate with an unstable oligomer (nucleus) that limits the rate of polymer formation; once formed, the polymer then elongates efficiently from this nucleus by monomer addition. Such models faithfully describe actin assembly, but they fail to account for how more complex polymers like hollow microtubules assemble. Here, we articulate a new model for microtubule formation that has three key features: (1) microtubules initiate via rectangular, sheet-like structures that grow faster the larger they become; (2) the dominant pathway proceeds via accretion, the stepwise addition of longitudinal or lateral layers; and (3) a “straightening penalty” to account for the energetic cost of tubulin’s curved-to-straight conformational transition. This model can quantitatively fit experimental assembly data, providing new insights into biochemical determinants and assembly pathways for microtubule nucleation. 

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