Ion specific effects in bundling and depolymerization of taxol-stabilized microtubules

Abstract

Microtubules (MTs) are nanometer scale hollow cylindrical biological polyelectrolytes. They are assembled from α/β-tubulin dimers, which stack to form protofilaments (PFs) with lateral interactions between PFs resulting in the curved MT. In cells, MTs and their assemblies are critical components in a range of functions from providing tracks for the transport of cargo to forming the spindle structure during mitosis. Previous studies have shown that while cations with valence equal to or larger than 3+ tend to assemble tight 3D bundles of taxol-stabilized MTs, certain divalent cations induce relatively loose 2D bundles of different symmetry [D. J. Needleman, et al., PNAS, 2004, 101, 16099]. Similarly, divalent cations form 2D bundles of DNA adsorbed on cationic membranes [I. Koltover, et al., PNAS, 2000, 97, 14046]. The bundling behavior for these biological polyelectrolyte systems is qualitatively in agreement with current theory. Here, we present results, which show that unlike the case for DNA adsorbed on cationic membranes, bundling of taxol-stabilized MTs occurs only for certain divalent cations above a critical ion concentration (e.g. Ca2+, Sr2+, Ba2+). Instead many divalent cations preempt the bundling transition and depolymerize taxol-stabilized MTs at a lower counterion concentration. Although previous cryogenic TEM has shown that, in the absence of taxol, Ca2+ depolymerizes MTs assembling in buffers containing GTP (guanosine triphosphate), our finding is surprising given the known stabilizing effects of taxol on GDP (guanosine diphosphate)-MTs. The ion concentration required for MT depolymerization decreases with increasing atomic number for the divalents Mg2+, Mn2+, Co2+, and Zn2+. GdCl3 (3+) is found to be extremely efficient at MT depolymerization requiring about 1 mM, while oligolysine (2+), is observed not to depolymerize MTs at concentrations as high as 144 mM. The surprising MT depolymerization results are discussed in the context of divalents disrupting either lateral interactions between PFs (which are strengthened for taxol containing β-tubulin) or interfering with taxol’s ability to induce flexibility at the interface between two tubulin dimers in the same PF (which has been recently suggested as a mechanism by which taxol stabilizes MTs post-hydrolysis with the induced flexibility counteracting the kink between GDP-tubulin dimers in a PF).

Self-assembly of biomolecular soft matter

“…Safinya and co-workers, for example, reported very interesting ion specific effects on depolymerization of taxol-stabilized microtubules. 101 It is fascinating that specific interactions with physiologically relevant ions can depolymerize these one-dimensional supramolecular structures even though they are stabilized by molecules well known to prevent their disassembly. Could there be similar strategies that massively disassemble amyloids?…”

The effect of multivalent cations and Tau on paclitaxel-stabilized microtubule assembly, disassembly, and structure

“…In this section we describe the finding that a significant number of divalent cations, notably, the biological ions Mg 2+ , Mn 2+ , and Zn 2+ , and Co 2+ (normally absent in vivo), do not bundle paclitaxel-stabilized MTs. The lack of MT bundling is substantiated in SAXS data, with scattering profiles consistent with the form factor of MTs modeled as nanotubes with no evidence of a bundling correlation peak [44]. Instead, above a critical ion concentration, the form factor is replaced by an enhanced and featureless SAXS scattering indicative of depolymerization of paclitaxel-stabilized MTs into curled protofilament rings (Fig.…”

“…4b. The y-axis is a plot of the MT radius (from fits to the SAXS data [44]) and the x-axis shows the range over which MTs are stable for each cation. For each ion plotted, the end of the color coded arrow corresponds to the highest ion concentration where SAXS data indicates stable MTs.…”

“…The y-axis shows that the MT inner radius (derived from fits to SAXS data) is unchanged in the presence of the different ions. Reproduced from [44] with permission from the Royal Society of Chemistry.…”

Trivalent Cation Induced Bundle Formation of Filamentous fd Phages

“…The role of counterions is to induce attractive forces between like‐charged rod‐shaped polyelectrolytes such as MT filaments and to hinder repulsive forces. Counterion‐mediated bundle formation is mainly influenced by the valence of the applied counterions which can be explained by Manning's theory of condensation . As MTs are exposed to counterions, they tend to condense on polyelectrolyte surfaces preventing repulsive forces between MT rods.…”