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Reversibility and Viscoelastic Properties of Micropillar Supported and Oriented Magnesium Bundled F-Actin.

Maier T, Haraszti T - PLoS ONE (2015)

Bottom Line: Understanding the bundling kinetics of actin filaments is important in the formation of various cytoskeletal structures, such as filopodia and stress fibers.Microparticles attached to the filaments allowed the measurement of thermal motion, and we found that bundling takes place at lower concentrations than previously found in 3-dimensional actin gels, i.e. actin filaments formed bundles in the presence of 5-12 mM of magnesium chloride in a time-dependent manner.The filaments also displayed long term stability for up to hours after removing the magnesium ions from the buffer, which suggests that there is an extensive hysteresis between cation induced crosslinking and decrosslinking.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute for Intelligent Systems, Department of New Materials and Biosystems, Heisenberg str. 3, D-70569 Stuttgart, Germany; University of Heidelberg, Institute of Physical Chemistry, Department of Biophysical Chemistry, Im Neuenheimer Feld 253, D-69120 Heidelberg, Germany.

ABSTRACT
Filamentous actin is one of the most important cytoskeletal elements. Not only is it responsible for the elastic properties of many cell types, but it also plays a vital role in cellular adhesion and motility. Understanding the bundling kinetics of actin filaments is important in the formation of various cytoskeletal structures, such as filopodia and stress fibers. Utilizing a unique pillar-structured microfluidic device, we investigated the time dependence of bundling kinetics of pillar supported free-standing actin filaments. Microparticles attached to the filaments allowed the measurement of thermal motion, and we found that bundling takes place at lower concentrations than previously found in 3-dimensional actin gels, i.e. actin filaments formed bundles in the presence of 5-12 mM of magnesium chloride in a time-dependent manner. The filaments also displayed long term stability for up to hours after removing the magnesium ions from the buffer, which suggests that there is an extensive hysteresis between cation induced crosslinking and decrosslinking.

No MeSH data available.


Related in: MedlinePlus

Flow channel for the creation of stress-fiber like actin structures.(A) A photograph of the prepared channel. (B) Schematic side view: The channel is imprinted into a thin PDMS layer on top of a cover slip (1) and closed with a ~3–4 mm thick block (2) with the pillar field (3) placed in the middle. The attached tubes (4) allow a continuous, pump driven exchange of liquid. (The scales are altered for better visibility.)
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pone.0136432.g001: Flow channel for the creation of stress-fiber like actin structures.(A) A photograph of the prepared channel. (B) Schematic side view: The channel is imprinted into a thin PDMS layer on top of a cover slip (1) and closed with a ~3–4 mm thick block (2) with the pillar field (3) placed in the middle. The attached tubes (4) allow a continuous, pump driven exchange of liquid. (The scales are altered for better visibility.)

Mentions: The flow channels (Fig 1) were constructed from two main parts: a main channel and a micropillar array protruding into the channel. The channels were typically 40 μm high, with a 6 mm × 6 mm main chamber in the middle. The pillars were 15 μm high with a diameter of 5 μm and a typical center-to-center distance of 13 μm. The pillar field was 5 mm × 5 mm in size, placed in the middle of the main chamber during assembly. Both parts were produced by standard soft lithography methods [27, 28] using SU-8 negative resists (MicroChem Corp., USA), treated according to the manufacturer’s instructions. Structures were exposed using a MJB3 mask aligner (SUESS MicroTec AG, Germany) equipped with a 350 W mercury lamp.


Reversibility and Viscoelastic Properties of Micropillar Supported and Oriented Magnesium Bundled F-Actin.

Maier T, Haraszti T - PLoS ONE (2015)

Flow channel for the creation of stress-fiber like actin structures.(A) A photograph of the prepared channel. (B) Schematic side view: The channel is imprinted into a thin PDMS layer on top of a cover slip (1) and closed with a ~3–4 mm thick block (2) with the pillar field (3) placed in the middle. The attached tubes (4) allow a continuous, pump driven exchange of liquid. (The scales are altered for better visibility.)
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4556452&req=5

pone.0136432.g001: Flow channel for the creation of stress-fiber like actin structures.(A) A photograph of the prepared channel. (B) Schematic side view: The channel is imprinted into a thin PDMS layer on top of a cover slip (1) and closed with a ~3–4 mm thick block (2) with the pillar field (3) placed in the middle. The attached tubes (4) allow a continuous, pump driven exchange of liquid. (The scales are altered for better visibility.)
Mentions: The flow channels (Fig 1) were constructed from two main parts: a main channel and a micropillar array protruding into the channel. The channels were typically 40 μm high, with a 6 mm × 6 mm main chamber in the middle. The pillars were 15 μm high with a diameter of 5 μm and a typical center-to-center distance of 13 μm. The pillar field was 5 mm × 5 mm in size, placed in the middle of the main chamber during assembly. Both parts were produced by standard soft lithography methods [27, 28] using SU-8 negative resists (MicroChem Corp., USA), treated according to the manufacturer’s instructions. Structures were exposed using a MJB3 mask aligner (SUESS MicroTec AG, Germany) equipped with a 350 W mercury lamp.

Bottom Line: Understanding the bundling kinetics of actin filaments is important in the formation of various cytoskeletal structures, such as filopodia and stress fibers.Microparticles attached to the filaments allowed the measurement of thermal motion, and we found that bundling takes place at lower concentrations than previously found in 3-dimensional actin gels, i.e. actin filaments formed bundles in the presence of 5-12 mM of magnesium chloride in a time-dependent manner.The filaments also displayed long term stability for up to hours after removing the magnesium ions from the buffer, which suggests that there is an extensive hysteresis between cation induced crosslinking and decrosslinking.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute for Intelligent Systems, Department of New Materials and Biosystems, Heisenberg str. 3, D-70569 Stuttgart, Germany; University of Heidelberg, Institute of Physical Chemistry, Department of Biophysical Chemistry, Im Neuenheimer Feld 253, D-69120 Heidelberg, Germany.

ABSTRACT
Filamentous actin is one of the most important cytoskeletal elements. Not only is it responsible for the elastic properties of many cell types, but it also plays a vital role in cellular adhesion and motility. Understanding the bundling kinetics of actin filaments is important in the formation of various cytoskeletal structures, such as filopodia and stress fibers. Utilizing a unique pillar-structured microfluidic device, we investigated the time dependence of bundling kinetics of pillar supported free-standing actin filaments. Microparticles attached to the filaments allowed the measurement of thermal motion, and we found that bundling takes place at lower concentrations than previously found in 3-dimensional actin gels, i.e. actin filaments formed bundles in the presence of 5-12 mM of magnesium chloride in a time-dependent manner. The filaments also displayed long term stability for up to hours after removing the magnesium ions from the buffer, which suggests that there is an extensive hysteresis between cation induced crosslinking and decrosslinking.

No MeSH data available.


Related in: MedlinePlus