Nanomechanical studies of vimentin intermediate filaments

Abstract

Intermediate filaments, microtubules and microfilaments are the major components of the cytoskeleton. Though it is known that intermediate filaments play an important role in the mechanical behaviour of cells, it is surprising that their mechanical properties are far from being fully understood.

The morphology and assembly process of the vimentin intermediate filaments (IFs) were studied using transmission electron microscopy (TEM) and atomic force spectroscopy (AFM). The width of the vimentin was found to change as the assembly proceeded. This finding agrees with the literature about the compaction process of vimentin IFs. The width of the IFs decreased gradually, while the range of width increased within the first few minutes after assembly initiation, and then decreased at last and became stable at 12.80±2.20nm. The average length of the IFs increased with decreasing rate. The length attained 485.60±162.23nm at 120 minutes. The range of length increased which revealed the assembly process was randomly occurring between filaments in the solution. The height of the IFs obtained with AFM did not show the periodicity in contrary to the literature. It may be due to the flattening of IFs on the functionalized mica(AP-mica) surface, or the periodicity was not prominent to be observed morphologically.

In the force spectroscopy study, the nanomechanical properties of individual vimentin intermediate filaments were studied using AFM. Fresh vimentin intermediate filaments and samples fixed with glutaraldehydewere examined, and force-displacement curves with nano-scale resolutions of different vimentin intermediate filament samples were analysed. The use of glutaraldehydefixative provided cross-linking of the IF, and the structural change will result in differences in their force-displacement curves which helped to provide comparison with the non-fixed samples in order to identify the structure-mechanical property relationship. Statistical studies of these curves revealed that tearing off of protofilaments from the mature intermediate filaments (with and without glutaraldehyde) occurred inthe low force regime below 100pN, and successive tearing off events were observed readily below 25 nm separations, which were comparable with the lengths of domains of around 20 nm. Different features of sawtooth indicated the possibility of sliding mechanism in vimentin IF, and the sliding was found to occur at 30.44±13.41pN. Helical domain unfoldings were observed only in the non-fixed samples to start at 10.19±5.63pN on average witha mean increase of 42.12±26.74nm. This force agreed with the prediction of the extended Bell model described in the literature and the length increase was around double of the domain length, which indicated the uncoiling of the coiled-coils. The force-displacement curves also reveal different modes of failure of the vimentin intermediate filaments including protofilaments slippage/sliding and entropic elasticity. A new tearing off model was hence proposed based on different modes of failure and a previous model developed for desmin filaments reported in literature.