Microtubule Gliding and Cross-Linked Microtubule Networks on Micropillar Interfaces

Abstract

We combined biochemical and topographical patterning to achieve motor-driven microtubule gliding on top of microfabricated pillar arrays with limited and controllable surface interactions of gliding microtubules. Kinesins immobilized on pillar heads pushed microtubules from the top of one micropillar to the next bridging up to 20 mum deep gaps filled with buffer solution. Distances of more than 10 mum were by-passed, and microtubule buckling was occasionally observed. The velocity distributions of microtubules gliding on poly(dimethylsiloxane) (PDMS) pillars, on flat PDMS, and on glass were found to be different, most likely due to topological and/or chemical differences between the substrates. We also used pillar arrays to suspend cross-linked microtubule networks, whose structural characteristics were governed by the topographical characteristics of the pillar pattern. These experiments open new possibilities to study the dynamics and the self-organization of motor/microtubule networks in defined topologically structured environments.

Early Keratinocyte Differentiation on Micropillar Interfaces

“…1 On the microscale, a new generation of model surfaces based on poly(dimethylsiloxane) (PDMS) pillars has allowed for studying the adaptation of the cytoskeleton to external mechanical and biochemical stimulations. 2 Moreover, biofunctionalization of discretized pillar surfaces has been employed for cell attachment. 3 These elastomeric model surfaces have a tremendous advantage in that they render micromechanical sensors for the measurement of mechanical interactions between cells and their extracellular environment.…”

“…For this purpose, the molds were emulsified with a homogeneous thick layer of photoreactive polymer. 20 One key to our experiments was to facilitate adhesion of IHGKs on the pillars. Because of its role in healing epithelial wounds and its use as a cell adhesion supporting molecule for many cell types, 21,22 we have chosen the ECM molecule FN for pillar biofunctionalization.…”

Elucidating the molecular mechanisms underlying cellular response to biophysical cues using synthetic biology approaches

“…This for instance allows studies of interactions between microtubules and kinesin molecular motors with only minimally perturbing environments. 80 Other examples include an assembled 2D network of microtubules on pillars, which was utilized in a combined fluorescence study to investigate the interaction between the kinesin motors themselves. 81 Using micropillars and super-resolution fluorescence microscopy it was discovered that there exists attractive interactions between kinesin motors, resulting in motor clustering along the microtubule as shown in Figure 5.…”

“…The combination of a functionalized flow cell and total internal reflection microscopy (TIRF) was utilized and it was demonstrated that microtubules can be guided by actin and conversely, microtubules can define actin growth and organization. 90,91 Many other in vitro systems can be used to reveal fundamental biophysical cytoskeletal properties, such as the dynamics of single actin filaments by microfluidics, 92 templated assembly of actomyosin bundles, 93 microtubule self-organization by the motor protein Eg5, 80 and the adhesion-driven spreading dynamics of biomimetic actin cortices. 94 …”

Micro- and Nanopatterning of Surfaces Employing Self Assembly of Nanoparticles and Its Application in Biotechnology and Biomedical Engineering

“…This results in regular arrays of nano-sized anchoring sites for biochemical functionalization via thiol chemistry. Control of spacing, order and biochemical function immobilized onto gold nanospots allowed for in depth studies of cellular functions (Spatz, Mössmer et al 2000;Roos, Ulmer et al 2005;Steinberg, Schulz et al 2006;Graeter, Huang et al 2007;Huang, Graeter et al 2009;Ranzinger, Krippner-Heidenreich et al 2009). …”