Mechanical Properties of Doubly Stabilized Microtubule Filaments

Abstract

Microtubules are cytoskeletal filaments responsible for cell morphology and intracellular organization. Their dynamical and mechanical properties are regulated through the nucleotide state of the tubulin dimers and the binding of drugs and/or microtubule-associated proteins. Interestingly, microtubule-stabilizing factors have differential effects on microtubule mechanics, but whether stabilizers have cumulative effects on mechanics or whether one effect dominates another is not clear. This is especially important for the chemotherapeutic drug Taxol, an important anticancer agent and the only known stabilizer that reduces the rigidity of microtubules. First, we ask whether Taxol will combine additively with another stabilizer or whether one stabilizer will dominate another. We call microtubules in the presence of Taxol and another stabilizer, doubly stabilized. Second, since Taxol is often added to a number of cell types for therapeutic purposes, it is important from a biomedical perspective to understand how Taxol added to these systems affects the mechanical properties in treated cells. To address these questions, we use the method of freely fluctuating filaments with our recently developed analysis technique of bootstrapping to determine the distribution of persistence lengths of a large population of microtubules treated with different stabilizers, including Taxol, guanosine-5' [(α, β)-methyleno] triphosphate, guanosine-5'-O-(3-thiotriphosphate), tau, and MAP4. We find that combinations of these stabilizers have novel effects on the mechanical properties of microtubules.

HIF Stabilization Weakens Primary Cilia

“…Fig 2 presents our results. As expected, addition of Taxol lowers the bending modulus in a concentration-dependent manner as reported elsewhere [32, 33]. Similarly, addition of CoCl 2 lowered the bending modulus, but it is not clear from our data if the action of CoCl 2 with respect to Taxol is independent or compensatory.…”

“…Because the action of Taxol on microtubule mechanics is well known [32, 33], we used Taxol as a control.…”

Effect of length and rigidity of microtubules on the size of ring-shaped assemblies obtained through active self-organization

“…23 Additionally, when compared to the rigidity of taxol stabilized GTP-MTs, rigidity of GMPCPP-MTs both in the presence and absence of taxol were reported to be higher. 31 In our experiments, ring-shaped MT assemblies obtained from taxol stabilized GTP-MTs were found to be smaller in size compared to that obtained from GMPCPP-MTs both in the presence and absence of taxol. was found no longer valid in our work where ~3.4 times difference in size of ring-shaped MT assembly was observed which is could be observed in the Figure 6.…”

Cytosolic Cl- Affects the Anticancer Activity of Paclitaxel in the Gastric Cancer Cell Line, MKN28 Cell

“…Briefly, tubulin stocks (10 mg/ml) were prepared by reconstituting purified porcine tubulin in a tubulin stock buffers (80 mM PIPES, 50 mM KCl, 2 mM MgCl 2 , 1 mM GTP or GTPγS (non-hydrolyzable analog of GTP) [22, 23], and 0.5 mM EGTA, pH 6.9 with KOH). Thereafter, tubulin stocks were diluted with normal Cl - glycerol buffer (40 mM PIPES, 50 mM KCl, 2 mM MgCl 2 , 5.6% Glycerol and 1 mM GTP or GTPγS) or low Cl - glycerol buffer (40 mM PIPES, 50 mM KNO 3 , 2 mM Mg(NO 3 ) 2 , 1 mM GTP or GTPγS, and 5.6% Glycerol) at various ratio to set final Cl - concentration were 5, 30, 45 and 50 mM with 10 µM PTX.…”