Lattice defects induced by microtubule-stabilizing agents exert a long-range effect on microtubule growth by promoting catastrophes

Abstract

Microtubules are dynamic cytoskeletal polymers that spontaneously switch between phases of growth and shrinkage. The probability of transitioning from growth to shrinkage, termed catastrophe, increases with microtubule age, but the underlying mechanisms are poorly understood. Here, we set out to test whether microtubule lattice defects formed during polymerization can affect growth at the plus end. To generate microtubules with lattice defects, we used microtubule-stabilizing agents that promote formation of polymers with different protofilament numbers. By employing different agents during nucleation of stable microtubule seeds and the subsequent polymerization phase, we could reproducibly induce switches in protofilament number and induce stable lattice defects. Such drug-induced defects led to frequent catastrophes, which were not observed when microtubules were grown in the same conditions but without a protofilament number mismatch. Microtubule severing at the site of the defect was sufficient to suppress catastrophes. We conclude that structural defects within the microtubule lattice can exert effects that can propagate over long distances and affect the dynamic state of the microtubule end.

Changes in seam number and location induce holes within microtubules assembled from porcine brain tubulin and in Xenopus egg cytoplasmic extracts

“…Microtubules with different numbers of seams have been described (Debs et al, 2020;des Georges et al, 2008;Howes et al, 2017;Kikkawa et al, 1994;Sosa et al, 1997), although it was not considered that both the seam number and location could vary within individual microtubules. Therefore, these previous studies and the present one indicate that the microtubule lattice is highly labile, with the ability to form different kinds of structural defects (Hunyadi et al, 2005;Rai et al, 2021).…”

CSPP1 stabilizes growing microtubule ends and damaged lattices from the luminal side

“…Importantly, there is also a notable difference between the effects microtubule-stabilizing drugs and CSPP1: taxanes induce structural defects (holes) in microtubule lattices because they promote switching in protofilament number (Rai et al, 2021). In contrast, CSPP1 seems to promote lattice integrity.…”

“…It is possible that the binding site of the CSPP1 MTB domain overlaps with that of Taxol, because we found some evidence of competition between Taxol and CSPP1 in microtubule stabilization assays. Importantly, there is also a notable difference between the effects microtubule-stabilizing drugs and CSPP1: taxanes induce structural defects (holes) in microtubule lattices because they promote switching in protofilament number (Rai et al, 2021). In contrast, CSPP1 seems to promote lattice integrity.…”

Dynamic microtubules slow down during their shrinkage phase

“…The change in depolymerization rate also implies that, similar to catastrophe, rescue may not be a single-step random process either and the probability of rescue may increase over time due to the slower shrinkage speed. Notably, recent studies have shown that repeated rescues can occur at distinct regions of the lattice, frequently associated with damage and structural irregularities of the lattice and their repair (24, 27, 31, 59, 60). Examining if the shrinkage rate also slows down at these rescue hotspots may indicate whether rescues promoted by these lattice irregularities or by rescue-promoting proteins share similar features.…”

“…A potential structural basis of multi-step catastrophe is the evolution of the tip structure, a form of memory (25, 26). For example, randomly occurring lattice defects, such as a change in protofilament number (27) or lattice-start number (28) may irreversibly alter the structural state of the growing tip making it more prone to catastrophe. Thus, length-dependent catastrophe, in addition to the growth speed fluctuations, also indicates that the growing tip changes its properties (i.e., state) over time.…”