Microtubule instability driven by longitudinal and lateral strain propagation

Abstract

Tubulin dimers associate longitudinally and laterally to form metastable microtubules (MTs). MT 1 disassembly is preceded by subtle structural changes in tubulin fueled by GTP hydrolysis. These 2 changes render the MT lattice unstable, but it is unclear how exactly they affect lattice energetics 3 and strain. We performed long-time atomistic simulations to interrogate the impacts of GTP 4 hydrolysis on tubulin lattice conformation, lateral inter-dimer interactions, and (non-)local lateral 5 coordination of dimer motions. The simulations suggest that most of the hydrolysis energy is 6 stored in the lattice in the form of longitudinal strain. While not significantly affecting lateral bond 7 stability, the stored elastic energy results in more strongly confined and correlated dynamics of 8 GDP-tubulins, thereby entropically destabilizing the MT lattice. 9 12 filamentous assemblies of αβ-tubulin dimers stacked head-to-tail in polar protofilaments (PFs) 13 and folded into hollow tubes via lateral interactions [1,2] (Fig. 1a). Each dimer binds two GTP 14 molecules of which only the one bound to β-tubulin is hydrolyzed in the MT lattice over time [3,4]. 15 This hydrolysis reaction is fundamental to MT dynamic instability [5], i.e. random switching 16 between phases of growth and shrinkage ( Fig. 1a). Remarkably, both slow assembly and rapid 17 disassembly of MTs -the latter termed catastrophe -are able to perform mechanical work because 18 each tubulin dimer is a storage of chemical energy [6][7][8]. 19 The switch from a relaxed 'curved' conformation of tubulin favored in solution to a higher-energy 20 'straight' one is inherent to MT assembly [9][10][11][12][13][14][15]. It allows growing MTs to recruit and temporarily 21 stabilize GTP-tubulin in the straight form, most likely due to the greater bending flexibility of PFs at intra-and inter-dimer interfaces [13,[16][17][18]. It is therefore conceivable that collapsing MTs 23 would follow a reverse pathway during disassembly; namely, they would release the conformational 24 tension stored in GDP-tubulins that lateral bonds can no longer counteract. However, due to 25 the system complexity and together with the inability of modern structural methods to directly 26 visualize all sequential steps in the GTPase cycle in the straight MT body at high resolutions, it is 27 still unknown exactly how and where the hydrolysis energy is converted to mechanical strain in the 28 lattice. 29Recent high-resolution cryo-EM studies have revealed, in line with the early finding [19], that 30 GTP hydrolysis triggers changes in α-tubulin, resulting in a subnanometer longitudinal lattice 31 compaction (Fig. 1b) [20-23]. Because by itself this global lattice rearrangement does not fully 32 indicate how it is linked to the strain accumulation at the single-dimer level, two competing scenarios 33 have been proposed [24]. According to the seam-centric or strain model of MT catastrophe [20-22], 34 1/17 the gradual build-up of longitudinal tension along the lattice upon GTP hydrolysis is the pri...

Bending-torsional elasticity and energetics of the plus-end microtubule tip

“…2C ). This finding correlates well with a recent MD study showing that GTP hydrolysis results in a ∼4 k B T longitudinal bond weakening [36] as well as with our previous finding that the longitudinal bonds in GDP-PFs are more fragile when subject to mechanical stretching [35]. We note, however, that the FDA provides only the enthalpic contribution to the longitudinal stability because the entropic penalty for eliminating solvent from tubulin's surface is not taken into account.…”

“…Relaxation dynamics and heterogeneity of MT plus-end tips At the MT tip, the elastic energy stored by straightened PFs is only insufficiently balanced by the total energy of lateral dimer-dimer interactions, which results in MT structures featuring irregularly splayed, curved PFs [25,26,27]. Recent atomistic MD simulation have predicted substantial differences in the bending rigidity of GTP-and GDP-bound free dimers, single PFs, and tubulin octamers [31,32,33,34,35,36]. Concomitantly, it has been proposed that the formation/breakage of lateral dimer-dimer contacts might also be nucleotide-dependent [37,38,39].…”

“…To this end, we constructed two initial atomistic models of GTP-and GDP-MT tips by consecutively stacking three identical MT segments, each comprising two layers of dimers and each optimized against the high-resolution cryo-EM densities of GMPCPP-or GDP-MTs [40,41,35] (see Fig. 1B and Methods for a detailed simulation protocol).…”

“…This stabilization arises mainly from rearrangements near the nucleotide binding pocket as well as from altered interactions between the H10 and H6 helices. Because the hydrolysis reaction does not happen at the intra-dimer interface [53], we speculate that the conformational changes triggered by GTP hydrolysis (lattice 'compaction' around the longitudinal inter-dimer interface [54,52,40,35]) propagate upstream, allosterically causing additional intra-dimer tensions.…”

Single depolymerizing and transport kinesins stabilize microtubule ends

“…Stabilization of this curvature is thought to weaken the bonds between terminal subunits (Arellano-Santoyo et al, 2017). Since microtubule protofilaments peel off during depolymerization, enhancing protofilament curvature and weakening lateral bonds between neighboring protofilaments can thus promote catastrophes (Igaev & Grubmüller, 2020;McIntosh et al, 2018). Indeed, while not all kinesin-8 motors have been shown to depolymerize microtubules, they do increase the catastrophe rate, but also promote rescues and, therefore, reduce the persistence length for both growth and shrinkage phases of dynamic microtubules (Du et al, 2010;Fukuda et al, 2014;Gardner, Zanic, Gell, Bormuth, & Howard, 2011;Su et al, 2011).…”