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Ewa Paluch

Cortical actomyosin gel breakage triggers shape oscillations in cells and cell fragments. Ewa Paluch. Present address: Max-Planck-Institute-CBG, Dresden. Institut Curie/CNRS, Paris. Cell crawling. [M. Abercrombie, Proc. R. Soc. Lond. B , 1980].

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Ewa Paluch

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  1. Cortical actomyosin gel breakage triggers shape oscillations in cells and cell fragments Ewa Paluch Present address: Max-Planck-Institute-CBG, Dresden Institut Curie/CNRS, Paris

  2. Cell crawling [M. Abercrombie, Proc. R. Soc. Lond. B, 1980] [B. Alberts et al., Molecular Biology of the Cell, 2002]

  3. Actin Myosin Microtubules Cell crawling - actin Cortex intrinsic dynamics??? nucleus [D. Bray, Cell Movements, 2001]

  4. Dynamics of the actomyosin cortex in suspension cells Microtubules L929 fibroblast

  5. Dynamics of the actomyosin cortex in suspension cells Microtubules PEG

  6. Dynamics of the actomyosin cortex in suspension cells Microtubules Nocodazole Microtubules PEG

  7. MT depolymerization Myosin II Regulatory Light Chain (RLC) GEF-H1 Rho Rho-kinase Myosin II Enhanced contractility Microtubules Nocodazole Microtubules PEG [B. Liu et al., Cell Adhes. Commun. 5:249-255 (1998)] [M. Krendel et al., Nat. Cell Biol. 4:294-301 (2002)]

  8. Microtubules Nocodazole Microtubules PEG Dynamics of the actomyosin cortex in suspension cells Lymphoblasts: [M. Bornens, M. Paintrand, C. Celati, J. Cell Biol. 109:1071-1083 (1989)]

  9. 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]

  10. 1. Actin and myosin II during the oscillation 2. The mechanism of the oscillation 3. Spontaneous cortical ruptures and cell motility

  11. 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]

  12. 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]

  13. 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]

  14. 1. The actomyosin shell breaks 2. A bulge is expelled and grows 1. Actin and myosin II during the oscillation Two-steps mechanism:

  15. 1. Actin and myosin II during the oscillation 2. The mechanism of the oscillation 3. Spontaneous cortical ruptures and cell motility

  16. Actin Myosin 2. The mechanism of the oscillation A mechanism for cortical symmetry breaking • The actin gel is under tension because of myosin motors • The integrated tension T is homogenous: σ σ Where the gel thinner, σ is higher. [K. Sekimoto et al., Eur. Phys. J. E, 13:247-259 (2004)]

  17. Actin (10% fluorescent) 2. The mechanism of the oscillation Beads mimicking the motility of Listeria monocytogenes Polystyrene beads coated with actin nucleator VCA Minimal mixture of proteins : Arp2/3, gelsolin, cofilin (+ profilin) (+ cross-linkers)

  18. Actin nucleator bead Actin gel 2. The mechanism of the oscillation Beads mimicking the motility of Listeria monocytogenes

  19. Stress R 2. The mechanism of the oscillation The actin gel growing around a bead undergoes stress because of the geometry [V. Noireaux et al., Biophys. J., 78:1643-1654 (2000)]

  20. 2. The mechanism of the oscillation Observation of symmetry breaking

  21. 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)]

  22. 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)]

  23. The controlled bead system can help understading cell cortex ruptures (role of various actin-binding proteins, etc). 2. The mechanism of the oscillation The actin shell around a bead: stress due to GEOMETRY Cell cortex: stress due to MYOSIN motors bead The rupture mechanism seems comparable.

  24. 2. The mechanism of the oscillation Bulge growth can be induced medium pipette: cytochalasin latrunculin… cell substrate

  25. pipette flow of medium cell 2. The mechanism of the oscillation Local stress application induces bulge growth P = 200 Pa [E. Paluch et al., Biophys. J., 89:724-733]

  26. 2. The mechanism of the oscillation Protrusion growth

  27. The oscillation is driven by contraction of the actomyosin cortex contraction velocity: 1 to 9 µm/min 2. The mechanism of the oscillation Is actomyosin shrinkage due to depolymerization or contraction?

  28. Actin Myosin 2. The mechanism of the oscillation A mechanism for the oscillation [E. Paluch, M. Piel, J. Prost, M. Bornens, C. Sykes, Biophys. J., 89:724-733]

  29. 1. Actin and myosin II during the oscillation 2. The mechanism of the oscillation 3. Spontaneous cortical ruptures and cell motility

  30. 3. Spontaneous cortical ruptures and cell motility Actin Myosin Microtubules … + microtubules reduce myosin II activity

  31. The oscillation and motility? [E. Paluch et al., Trends Cell Biol., in press]

  32. 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…” [D. J. Solecki et al., Nat. Neurosc., 7, 1195-1203 (2004)] Movie: http://www.nature.com/neuro/journal/v7/n11/extref/nn1332-S8.mpg

  33. 3. Spontaneous cortical ruptures and cell motility And if contractility is enhanced?

  34. 3. Spontaneous cortical ruptures and cell motility I. Contraction waves [K. Wolf et al., J. Cell Biol., 160:267-277 (2003)] II. Multipleblebs [E. Sahai, C. Marshall, Nat. Cell Biol., 5:711-719 (2003)] And if contractility is enhanced?

  35. 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]

  36. 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

  37. Physics group Cécile Sykes Jasper van der Gucht Julie Plastino Biology group Michel Bornens Matthieu Piel Theorists Jean-François Joanny Jacques Prost • GFP constructs: Beat Imhof (University of Geneva) • Rex Chisholm (Northwestern University, Chicago) • - Deconvolution: Jean-Baptiste Sibarita (Institut Curie)

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