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Cortical actomyosin gel breakage triggers
shape oscillations in cells and cell fragments
Ewa Paluch
Institut Curie/CNRS, Paris
Present address:
Max-Planck-Institute-CBG, Dresden
Cell crawling
[M. Abercrombie, Proc. R. Soc. Lond. B, 1980]
[B. Alberts et al., Molecular Biology of the Cell, 2002]
Cell crawling - actin
Cortex nucleus
intrinsic dynamics???
[D. Bray, Cell Movements, 2001]
Actin
Myosin
Microtubules
Dynamics of the actomyosin cortex in
suspension cells
Microtubules
L929 fibroblast
Dynamics of the actomyosin cortex in
suspension cells
Microtubules
PEG
Dynamics of the actomyosin cortex in
suspension cells
Microtubules
Nocodazole
PEG
Microtubules
MT depolymerization
GEF-H1
Rho
Rho-kinase
Myosin II Regulatory
Light Chain (RLC)
Myosin II
Enhanced contractility
[B. Liu et al., Cell Adhes. Commun. 5:249-255 (1998)]
[M. Krendel et al., Nat. Cell Biol. 4:294-301 (2002)]
Microtubules
Nocodazole
PEG
Microtubules
Dynamics of the actomyosin cortex in
suspension cells
Microtubules
Nocodazole
Microtubules
PEG
Lymphoblasts: [M. Bornens, M. Paintrand, C. Celati, J. Cell Biol. 109:1071-1083 (1989)]
Fragments of L929 fibroblasts
Movie: http://www.biophysj.org/content/vol0/issue2005/images/data/biophysj.105.060590/DC1/Paluch-Movie1.mov
Cytoplast
Centrifugation after
microfilaments and
microtubules
depolymerization
Fragments
L929 fibroblasts
Nucleus
[E. Paluch, M. Piel, J. Prost, M. Bornens, C. Sykes, Biophys. J., 89:724-733]
1. Actin and myosin II during the oscillation
2. The mechanism of the oscillation
3. Spontaneous cortical ruptures and cell motility
1. Actin and myosin II during the oscillation
Dynamic characterization of actin during the oscillation
Movie: http://www.biophysj.org/content/vol0/issue2005/images/data/biophysj.105.060590/DC1/Paluch-Movie3.mov
Cell fragment
5 µm
[E. Paluch et al., Biophys. J., 89:724-733]
1. Actin and myosin II during the oscillation
Dynamic characterization of actin during the oscillation
5 µm
[E. Paluch et al., Biophys. J., 89:724-733]
1. Actin and myosin II during the oscillation
Dynamic characterization of myosin II
during the oscillation
Movie: http://www.biophysj.org/content/vol0/issue2005/images/data/biophysj.105.060590/DC1/Paluch-Movie4.mov
5 µm
[E. Paluch et al., Biophys. J., 89:724-733]
1. Actin and myosin II during the oscillation
Two-steps mechanism:
1. The actomyosin shell breaks
2. A bulge is expelled and grows
1. Actin and myosin II during the oscillation
2. The mechanism of the oscillation
3. Spontaneous cortical ruptures and cell motility
2. The mechanism of the oscillation
A mechanism for cortical symmetry breaking
σ
σ
- The actin gel is under tension
because of myosin motors
- The integrated tension T is
homogenous:
rext
T   de
rint
Where the gel thinner,
σ is higher.
Actin
Myosin
[K. Sekimoto et al., Eur. Phys. J. E, 13:247-259 (2004)]
2. The mechanism of the oscillation
Beads mimicking the motility of
Listeria monocytogenes
Polystyrene beads coated
with actin nucleator VCA
Actin
(10% fluorescent)
Minimal mixture of proteins :
Arp2/3, gelsolin, cofilin (+ profilin)
(+ cross-linkers)
2. The mechanism of the oscillation
Beads mimicking the motility of
Listeria monocytogenes
Actin nucleator
bead
Actin gel
2. The mechanism of the oscillation
The actin gel growing around a bead
undergoes stress because of the geometry
Stress
R
[V. Noireaux et al., Biophys. J., 78:1643-1654 (2000)]
2. The mechanism of the oscillation
Observation of symmetry breaking
2. The mechanism of the oscillation
Symmetry breaking can be induced
Local photolysis of the actin gel
[J. van der Gucht, E. Paluch, J. Plastino, C. Sykes, PNAS, 22:7847-7852 (2005)]
2. The mechanism of the oscillation
Hole formation is reminiscent of a
fracture
2 µm
t = 13’
t = 15’
t = 17’
[J. van der Gucht, E. Paluch, J. Plastino, C. Sykes, PNAS, 22:7847-7852 (2005)]
2. The mechanism of the oscillation
Cell cortex: stress due to MYOSIN
motors
The actin shell around a bead:
stress due to GEOMETRY
bead
The rupture mechanism seems comparable.
The controlled bead system can help understading cell
cortex ruptures (role of various actin-binding proteins, etc).
2. The mechanism of the oscillation
Bulge growth can be induced
medium
cell
substrate
pipette: cytochalasin
latrunculin…
2. The mechanism of the oscillation
Local stress application induces bulge growth
pipette
cell
flow of medium
P = 200 Pa
[E. Paluch et al., Biophys. J., 89:724-733]
2. The mechanism of the oscillation
Protrusion growth
2. The mechanism of the oscillation
Is actomyosin shrinkage due to depolymerization or contraction?
contraction velocity:
1 to 9 µm/min
The oscillation is driven by contraction of the actomyosin cortex
2. The mechanism of the oscillation
A mechanism for the oscillation
Actin
Myosin
[E. Paluch, M. Piel, J. Prost, M. Bornens, C. Sykes, Biophys. J., 89:724-733]
1. Actin and myosin II during the oscillation
2. The mechanism of the oscillation
3. Spontaneous cortical ruptures and cell motility
3. Spontaneous cortical ruptures and cell motility
… + microtubules
reduce myosin II activity
Actin
Myosin
Microtubules
The oscillation and motility?
[E. Paluch et al.,
Trends Cell Biol., in press]
The oscillation and motility?
“Migration involves the coordinated two-stroke
movement of a perinuclear tubulin ‘cage’, and
the centrosome, with the centrosome moving
forward before nuclear translocation…”
QuickTime™ and a
YUV420 codec decompressor
are needed to see this picture.
[D. J. Solecki et al., Nat. Neurosc., 7, 1195-1203 (2004)]
Movie: http://www.nature.com/neuro/journal/v7/n11/extref/nn1332-S8.mpg
3. Spontaneous cortical ruptures and cell motility
And if contractility is enhanced?
3. Spontaneous cortical ruptures and cell motility
And if contractility is enhanced?
I. Contraction waves
[K. Wolf et al., J. Cell Biol., 160:267-277 (2003)]
II. Multiple blebs
[E. Sahai, C. Marshall, Nat. Cell Biol., 5:711-719 (2003)]
3. Spontaneous cortical ruptures and cell motility
And if contractility is enhanced?
- Blebs result from cortical
contractility
- Dual behavior comparable to
e.g. growth of dry zones in a thin
water film
revue:
[E. Paluch, C. Sykes, J. Prost, M. Bornens,
Dynamic modes of the cortical actomyosin
gel during cell locomotion and division,
Trends in Cell Biol., in press]
Summary
• Cortical oscillation is a general phenomenon resulting
from elastic gel properties of the actomyosin cortex
• Bleb formation reveals the level of cortical contractility
• Spontaneous cortical ruptures (and blebs) can be used by
cells or remain a side-product of cortex contractions
• Cortex breakage in cells // symmetry breaking of gels
around beads
Physics group
Cécile Sykes
Jasper van der Gucht
Biology group
Julie Plastino
Theorists
Michel Bornens
Matthieu Piel
Jean-François Joanny
Jacques Prost
- GFP constructs: Beat Imhof (University of Geneva)
Rex Chisholm (Northwestern University, Chicago)
- Deconvolution: Jean-Baptiste Sibarita (Institut Curie)