The main goal of the group’s research is to understand how the mechanical properties of the cell are regulated at the protein level to achieve controlled cellular deformations. To that aim, we study the cell cortex, a network of actin, myosin, and associated proteins that lies beneath the plasma membrane and determines the shape of the cell body. The cortex enables the cell to resist externally applied forces and exert mechanical work. As such, it plays a role in normal physiology during events that involve cell deformation, such as mitosis, cytokinesis, and cell locomotion, and in the pathophysiology of diseases, such as cancer, in which cortical contractility is often upregulated. Despite its importance, very little is known about how the cortex is assembled and regulated.
The biological function of the cortex relies on its ability to contract and exert forces. Therefore, the biological properties of the cortex cannot be understood in isolation from its mechanics. Our main focus is on investigating how cortical mechanical properties are determined by the molecular components of the cortex and how these properties are regulated, locally and globally, to allow the cell to undergo deformations during cell division and migration.
We are particularly interested in blebs, spherical membrane protrusions driven by contractions of the actomyosin cortex. Although blebs are commonly observed during apoptosis, cell spreading, cytokinesis, and migration, their growth and physiological functions are still poorly understood. We investigate the physical and biological mechanisms of bleb formation and study their function during cytokinesis and migration. Moreover, de novo cortex assembly can be observed at the bleb membrane, and we use blebs as tools for the study of cortex nucleation and growth. The staff, composed of biologists and physicists, combines biophysical and molecular approaches. Our main lines of research are the following:
1. Regulation of cortex assembly and cortex mechanics
Our aim is to understand the mechanisms and regulation of cortex assembly and steady-state turnover. Despite the physiological importance of the cortex, basic properties, such as cortex thickness, the spatial organization of the network, and its dynamical behavior (turnover), are very poorly understood. One reason for this is that the thickness of the cortical network is less than 1 μm, which makes it difficult to observe using conventional optical microscopy. We have developed a method of measuring cortex thickness and monitoring the dynamics of cortex turnover with sufficient spatial and temporal resolution. We are also investigating the de novo assembly of the cortex using cellular blebs as a model system. Indeed, blebs are initially devoid of filamentous actin and reassemble a contractile cortex prior to retraction. Thus, they present an ideal system for the study of cortex growth.
Figure 1: (A) Animal view of the leading edge of a wildtype zebrafish prechordal plate. The inset shows a schematic animal view of an embryo at 80% epiboly, with the green rectangle marking the imaged area. Examples of a bleb, filopodium, and lamellipodium in prechordal plate leading edge cells are shown. Arrowheads indicate protrusions. The arrow indicates the separation between the actin cortex and membrane in the bleb. Scale bars = 10 μm. (B) Percentage of blebs, filopodia, and lamellipodia in wildtype prechordal plate leading edge cells. (C) Lateral view of a MZoep mutant embryo (blue) with transplanted ERM-deficient mesendoderm cells (red). ERM-deficient cells form more blebs than control (green) cells and display a less straight migration path (compare the red and green tracks). Tracking time = 110 min. Scale bar = 50 μm. (Author: Alba Diz-Muñoz, figure based on Diz-Muñoz et al., PLoS Biol, 2010)
3. Mechanics of the dividing cell: cortex tension in cytokinesis
The formation and ingression of the cleavage furrow during cytokinesis relies on the controlled reorganization of the actin cortex. So far, most studies of cytokinetic mechanics have focused on force generation at the cell equator, where cortical actin and myosin accumulate in a contractile ring. However, a significant amount of actin and myosin is also present at the poles of a dividing cell.
Mechanical resistance of this polar cortex has been shown to slow ingression dynamics. We investigated the contribution of contractile forces exerted at the poles to cytokinesis and showed that polar contractility makes the symmetric shape of the cytokinetic cell intrinsically unstable. Indeed, an imbalance in contractile forces between the two poles can displace the cleavage furrow from its equatorial position, leading to shape instabilities that can result in division failure.
We showed that such instabilities are present during control division and can be amplified by treatments that affect the actin cortex, leading to shape oscillations and division failure. We have further shown that blebs, which are commonly observed at the poles of dividing cells, can prevent the appearance of shape instabilities by constantly releasing polar contractility. We are currently working on a physical model of the shape instabilities.
4. Bleb and lamellipodia during cell migration in threedimensional environments
In three-dimensional environments, bleb-based migration is a widespread alternative to lamellipodial migration and is commonly used by cancer cells and during development (Charras and Paluch, Nat Rev Mol Cell Biol, 2008). What determines the type of protrusion formed by a migrating cell and how the various protrusion types contribute to cell migration are poorly understood. We investigate the formation and function of blebs and lamellipodia in two different systems:
– We study cell migration in vivo during Danio rerio (zebrafish) embryonic development (collaboration with the laboratory of Prof. C.P. Heisenberg, IST, Austria). We showed that mesendoderm progenitor cells in the zebrafish prechordal plate migrate during gastrulation using a combination of blebs, lamellipodia, and filopodia. Therefore, they constitute an ideal system for investigating the respective contributions of the different protrusion types to cell migration. We have used a variety of methods to increase the proportion of blebs at the expense of the other protrusion types and showed that increasing bleb formation slows migration by reducing the directional persistence of the migrating cells (Fig. 1, Diz-Muñoz et al., PLoS Biol, 2010). We are currently investigating the effects of decreasing bleb or enhancing lamellipodia formation on mesendoderm progenitor migration. We are also characterizing the orientation of the various protrusion types with respect to the migration direction.
- In parallel, we are investigating the mechanisms of bleb and lamellipodia formation using an in vitro culture system. We use Walker carcinosarcoma cells, which can be induced to form either blebs or lamellipodia, depending on the culture conditions. We are currently comparing the mechanical properties of the two sublines and the molecular pathways that lead to the formation of one or the other protrusion type.