In in vivo situation, the imbalance in the osmotic pressure will inflate the cell
resulting in the increased membrane tension, and also various other loads will be
exerted against the protrusion. For example, immune cells must crawl through the
cells constituting the blood vessel wall. Penetrating the membrane protrusion into
the space between cells will impose a considerable resistance against the protrusion.
The co-existence of the fluctuation of the cell membrane and actin filaments will be
advantageous under these circumstances.
7.19 Reconstituted Systems to Study the PolymerizationBased Phenomena
7.19.1 Actin-Liposome System
In this section, mimicking the growth of the protrusions of the cell edge by using an
actin-encapsulating liposome is described. Two studies appeared in the early stage of
the research. In one study [178], micron-sized liposomes containing actin monomer
were formed by swelling the lipid film in a solution containing actin monomer. The
actin-encapsulating liposomes were observed under light microscope. The actin
monomer in liposomes was polymerized by K
+ ions that were introduced using an
ionophore. Polymerization of encapsulated actin was confirmed by fluorescence
photo- bleaching recovery (FPR; Sect. 3.6.2) of the actin that had been fluorescently
labeled with rhodamine. As a result of polymerization, the liposome assumed
irregular shapes. Actin crosslinking protein filamin altered the final shape of the
liposome to less irregular shapes suggesting that the lipid membranes could be
sculpted by the structures formed by the combination of actin filaments and actin
crosslinking proteins.
Another study [36] also observed actin-containing liposomes, but showing the
process of morphological changes. Actin was allowed to slowly polymerize with a
low concentration of Ca
2+ ions that had been co-encapsulated on ice with actin
monomers. Polymerization was initiated by raising temperature from 5
C to 30
C.
With the progress of the polymerization, the shape of the liposome changed from
spherical to a disk- or a racket-like shape. In both types of liposomes, birefringence
was observed along the contour of the deformed liposomes, which was interpreted as
an alignment of actin filaments along the liposome contour. The shape change
occurred slowly over several tens of minutes concomitant with the actin polymerization (the actin polymerization was separately confirmed by an increase in the light
scattering of the actin solution).
The time scale of the above experiment was much longer than the time scale of
the dynamics of lamellipodia (several tens of seconds in amoeba cell and in
mammalian cells). Therefore, a method was developed that allowed faster polymerization [179]. To accelerate polymerization, polymerization nuclei (fragments of
actin filaments) were incorporated with actin monomer. But most importantly,
140
7 Moving Life
resulting in the increased membrane tension, and also various other loads will be
exerted against the protrusion. For example, immune cells must crawl through the
cells constituting the blood vessel wall. Penetrating the membrane protrusion into
the space between cells will impose a considerable resistance against the protrusion.
The co-existence of the fluctuation of the cell membrane and actin filaments will be
advantageous under these circumstances.
7.19 Reconstituted Systems to Study the PolymerizationBased Phenomena
7.19.1 Actin-Liposome System
In this section, mimicking the growth of the protrusions of the cell edge by using an
actin-encapsulating liposome is described. Two studies appeared in the early stage of
the research. In one study [178], micron-sized liposomes containing actin monomer
were formed by swelling the lipid film in a solution containing actin monomer. The
actin-encapsulating liposomes were observed under light microscope. The actin
monomer in liposomes was polymerized by K
+ ions that were introduced using an
ionophore. Polymerization of encapsulated actin was confirmed by fluorescence
photo- bleaching recovery (FPR; Sect. 3.6.2) of the actin that had been fluorescently
labeled with rhodamine. As a result of polymerization, the liposome assumed
irregular shapes. Actin crosslinking protein filamin altered the final shape of the
liposome to less irregular shapes suggesting that the lipid membranes could be
sculpted by the structures formed by the combination of actin filaments and actin
crosslinking proteins.
Another study [36] also observed actin-containing liposomes, but showing the
process of morphological changes. Actin was allowed to slowly polymerize with a
low concentration of Ca
2+ ions that had been co-encapsulated on ice with actin
monomers. Polymerization was initiated by raising temperature from 5
C to 30
C.
With the progress of the polymerization, the shape of the liposome changed from
spherical to a disk- or a racket-like shape. In both types of liposomes, birefringence
was observed along the contour of the deformed liposomes, which was interpreted as
an alignment of actin filaments along the liposome contour. The shape change
occurred slowly over several tens of minutes concomitant with the actin polymerization (the actin polymerization was separately confirmed by an increase in the light
scattering of the actin solution).
The time scale of the above experiment was much longer than the time scale of
the dynamics of lamellipodia (several tens of seconds in amoeba cell and in
mammalian cells). Therefore, a method was developed that allowed faster polymerization [179]. To accelerate polymerization, polymerization nuclei (fragments of
actin filaments) were incorporated with actin monomer. But most importantly,
140
7 Moving Life