formed by bridging the actomyosin bundle grown from the focal adhesion to the
actin bundles formed beneath the upper surface of the cell [104, 109, 110]. Formation
of the bundle structure of filaments with opposing orientation is achieved by alphaactinin.
The focal adhesion bears the tension generated by the stress fiber. Interestingly,
this tension is necessary for the maintenance of the stress fiber itself [111]. This
interesting property has led to a suggestion that the contractile force acting within the
actin network gathers actin filaments [112]. In a number of studies (eg., [113]),
application of a uni-axial force to the cell aligns actin filaments along the force axis
to generate a fibrous bundle-like actin structure. The decreased ATPase activity
(corresponding to the contractile force) of myosin II by inhibitors resulted in the loss
of fibrous structure, whereas removal of inhibitors resulted in the re-appearance of
the fibrous structures. Thus, the external force plays important roles in the stress fiber
maintenance and formation. In particular, it enhances the linkage of actin filament to
integrin cluster; talin, an actin-crosslinking protein, makes a weak bond with actin
filament, but under the external force, it undergoes a conformational change and
becomes able to bind to another protein, vinculin, that also binds to actin filament,
establishing more stable contact between the actin filament and the integrin cluster
([114] and references therein). Second, the external force recruits another protein,
zyxin, to focal adhesion and in the middle of the stress fiber [115]. It is hypothesized
that zyxin is activated by the external force and then recruits a protein, Ena/VASP, to
the focal adhesion and to the middle of stress fiber. Ena/VASP mediates the
extracellular signal to nucleation of actin polymerization by binding to both receptor
and the nucleator for the polymerization [116]. For the sensing of the external force,
mechano-sensing protein(s) has been postulated to localize to stress fiber and focal
adhesion to which zyxin is bound [115].
Contraction-competent stress fiber has been isolated [104], indicating that the
stress fiber is an independent entity. On the other hand, it has been shown that
fluorescently labeled actin is rapidly incorporated into pre-existing stress fiber [117]
and that fluorescently-labeled alpha-actinin constantly exchanged [118]. The rapid
turnover of actin in stress fiber and the stability of the stress fiber seem to be difficult
to reconcile, but the latter may be because of the use of low ionic strength buffer
devoid of ATP, under which actin-myosin interaction is strong and stabilized the
isolated bundle structure.
7.12 Actin Filaments in Lamellipodium and Lamella
Fluorescent staining of cellular actin demonstrated that there are two major staining
patterns in the cell: uniform staining that is prominent in lamellipodia and streaks
penetrating through the uniform staining (Fig. 7.19 and 7.21a). The former is
thought to represent an actin filament network in lamellipodium and the latter the
stress fiber/bundle continuing to the filopodium. Intracellular myosin II visualized by
immunofluorescence microscopy, as shown in Fig. 7.21b, is found to distribute in
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7 Moving Life
actin bundles formed beneath the upper surface of the cell [104, 109, 110]. Formation
of the bundle structure of filaments with opposing orientation is achieved by alphaactinin.
The focal adhesion bears the tension generated by the stress fiber. Interestingly,
this tension is necessary for the maintenance of the stress fiber itself [111]. This
interesting property has led to a suggestion that the contractile force acting within the
actin network gathers actin filaments [112]. In a number of studies (eg., [113]),
application of a uni-axial force to the cell aligns actin filaments along the force axis
to generate a fibrous bundle-like actin structure. The decreased ATPase activity
(corresponding to the contractile force) of myosin II by inhibitors resulted in the loss
of fibrous structure, whereas removal of inhibitors resulted in the re-appearance of
the fibrous structures. Thus, the external force plays important roles in the stress fiber
maintenance and formation. In particular, it enhances the linkage of actin filament to
integrin cluster; talin, an actin-crosslinking protein, makes a weak bond with actin
filament, but under the external force, it undergoes a conformational change and
becomes able to bind to another protein, vinculin, that also binds to actin filament,
establishing more stable contact between the actin filament and the integrin cluster
([114] and references therein). Second, the external force recruits another protein,
zyxin, to focal adhesion and in the middle of the stress fiber [115]. It is hypothesized
that zyxin is activated by the external force and then recruits a protein, Ena/VASP, to
the focal adhesion and to the middle of stress fiber. Ena/VASP mediates the
extracellular signal to nucleation of actin polymerization by binding to both receptor
and the nucleator for the polymerization [116]. For the sensing of the external force,
mechano-sensing protein(s) has been postulated to localize to stress fiber and focal
adhesion to which zyxin is bound [115].
Contraction-competent stress fiber has been isolated [104], indicating that the
stress fiber is an independent entity. On the other hand, it has been shown that
fluorescently labeled actin is rapidly incorporated into pre-existing stress fiber [117]
and that fluorescently-labeled alpha-actinin constantly exchanged [118]. The rapid
turnover of actin in stress fiber and the stability of the stress fiber seem to be difficult
to reconcile, but the latter may be because of the use of low ionic strength buffer
devoid of ATP, under which actin-myosin interaction is strong and stabilized the
isolated bundle structure.
7.12 Actin Filaments in Lamellipodium and Lamella
Fluorescent staining of cellular actin demonstrated that there are two major staining
patterns in the cell: uniform staining that is prominent in lamellipodia and streaks
penetrating through the uniform staining (Fig. 7.19 and 7.21a). The former is
thought to represent an actin filament network in lamellipodium and the latter the
stress fiber/bundle continuing to the filopodium. Intracellular myosin II visualized by
immunofluorescence microscopy, as shown in Fig. 7.21b, is found to distribute in
126
7 Moving Life