Atomic force microscopy reveals distinct protofilament-scale structural dynamics in depolymerizing microtubule arrays

The dynamic reorganization of microtubule-based cellular structures, such as the spindle and the axoneme, fundamentally depends on the dynamics of individual polymers within multimicrotubule arrays. A major class of enzymes implicated in both the complete demolition and fine size control of microtubule-based arrays are depolymerizing kinesins. How different depolymerases differently remodel microtubule arrays is poorly understood. A major technical challenge in addressing this question is that existing optical or electron-microscopy methods lack the spatial-temporal resolution to observe the dynamics of individual microtubules within larger arrays. Here, we use atomic force microscopy (AFM) to image depolymerizing arrays at single-microtubule and protofilament resolution. We discover previously unseen modes of microtubule array destabilization by conserved depolymerases. We find that the kinesin-13 MCAK mediates asynchronous protofilament depolymerization and lattice-defect propagation, whereas the kinesin8 Kip3p promotes synchronous protofilament depolymerization. Unexpectedly, MCAK can depolymerize the highly stable axonemal doublets, but Kip3p cannot. We propose that distinct protofilament-level activities underlie the functional dichotomy of depolymerases, resulting in either large-scale destabilization or length regulation of microtubule arrays. Our work establishes AFM as a powerful strategy to visualize microtubule dynamics within arrays and reveals how nanometer-scale substrate specificity leads to differential remodeling of micron-scale cytoskeletal structures.

Automated summary

This study uses atomic force microscopy (AFM) to observe the dynamic reorganization of microtubule arrays by depolymerizing enzymes. Previously unseen destabilization modes of microtubule arrays by depolymerases are discovered. The study reveals that different depolymerases exhibit distinct depolymerization patterns, leading to either large-scale destabilization or length regulation of microtubule arrays.