Physical and Molecular Principles Governing Cytoskeletal Organization
Group leader : A. Michelot
Our team has two main objectives. Our first aim is to understand some of the basic physical and biochemical principles governing cytoskeletal organization (principally actin) in eukaryotic cells. Our second aim is to understand how cells use different actin networks to perform a variety of cellular functions.
The transition of a tumor to malignancy is associated with many genetic and epigenetic modifications of its cells. For some cancers, it is established that part of these modifications corresponds to changes in the expression level of some proteins implicated in the regulation of the actin cytoskeleton. The actin cytoskeleton is playing a major role in controlling cells’ structural integrity and in their dynamics within tissues. As a consequence, cancer cells have altered motile and biomechanical properties, but these properties are also well adapted for their propagation during the formation of metastases.
The actin cytoskeleton is composed of dense filamentous networks. These networks are essential for a large number of cellular processes implicating the generation of forces, or the resistance to mechanical constraints, such as cell motility, adhesion or division. The reason why actin-based structures, all composed of filaments assembled from identical subunits, are able to perform many different functions is due to the fact that cells are able to organize actin filaments in a wide variety of structures. Each of these structures has specific geometrical, dynamical and rheological properties that are adapted for a given cellular process. The properties of each individual networks are constantly remodeled by specific sets of actin binding proteins (ABPs), that are responsible for the nucleation, crosslinking, and/or disassembly of the filaments.
In this context, our objective is to determine how cells can generate a variety of structures of actin filaments with defined properties, and with appropriate protein composition, from a common pool of cytoplasmic components. This global understanding will enable us to predict how misbalancing the equilibrium of the cell affects actin networks organizational, dynamical and mechanical properties.
Self-organization of the actin cytoskeleton
To develop a working model, we are left with the complex question of what type of signaling may cells use to target precisely, spatially and temporally, multiple families of ABPs to the appropriate actin networks. Previously, we demonstrated that the local recruitment of specific actin nucleation promoting factor is sufficient to assemble from cell extracts actin networks of appropriate protein composition. This surprising observation indicates that recruitment of ABPs is not due to a specific signaling network, or as a local cellular context, such as pH or salt concentration, but on the contrary that the segregation of ABPs to appropriate structures is due to a fine-tuned biochemical regulation, although the underlying molecular mechanisms remain largely unknown.
Key regulators of actin filaments identity
Hence, what distinguishes different populations of actin filaments, although they are all assembled from identical actin subunits? In our projects, we are exploring several hypothesis to understand how identical actin filaments acquire a specific identity. In particular, recent research suggests that many proteins characterized previously as actin-filament bundling proteins may have an important function in giving actin filaments their identity. The decoration of actin filaments by subsets of actin binding protein may influence strongly the binding of others, through competitive or collaborative effects.
Inter-dependence of actin networks
More and more data indicate that actin networks in cells are in homeostasis. In other words, it means that actin networks compete for the same pool of ABPs, and that a perturbation of one actin network will disturb the whole equilibrium and affect also other actin networks. For example, the absence of an actin binding protein can not only affect the structural integrity of the actin structure where it normally binds to, but we also have evidences that it can modify the localization of other actin binding proteins, to trigger defects on other actin structures.
Our tools in the lab
To address detailed mechanistic questions, traditional biochemical, cell biological and genetic experiments need to be complemented by engineering-inspired reconstitution approaches. The use of simplified systems and the ability to modify individual components without the active contribution and complexity of the cell has proven beneficial in understanding emerging properties of a variety of biological systems.
We usually use two complementary ways to reconstitute our biological processes of interest. With “bottom-up” reconstitutions, we purify the essential components, and assemble them in a test tube. Components are added one-by-one into the system, and it allows for testing the role of a molecule in the presence of a limited number of partners. However, for cellular processes involving the participation of large numbers of families of molecules, “bottom-up” approaches can be limited, because they require the identification of all essential components and their purification in an active form. In that situation, we use protein extracts, because they contain a mixture of all the proteins that are essential for the process. We are experts in genetic depletions in protein extracts, in order to experiment “top-down” approaches, where components are removed one-by-one from the extracts, in order to test for their functions in a near-physiological environment. Both “bottom-up” and “top-down” approaches are used in various experimental setups to bridge the gap between simple biochemistry and the complexity of a cell.
January 28th, 2013
Actin filament elongation in Arp2/3-derived networks is controlled by three distinct mechanisms
January 21st, 2012
Actin cytoskeleton: a team effort during actin assembly
July 26th, 2011
Building distinct actin filament networks in a common cytoplasm
November 9th, 2010
Reconstitution and protein composition analysis of endocytic actin patches
May 15th, 2007
Actin-filament stochastic dynamics mediated by ADF/cofilin.
October 10th, 2006
A novel mechanism for the formation of actin-filament bundles by a nonprocessive formin.
August 17th, 2006
The formin homology 1 domain modulates the actin nucleation and bundling activity of Arabidopsis FORMIN1.
April 8th, 2013
Lsb1 is a negative regulator of las17 dependent actin polymerization involved in endocytosis.
November 22nd, 2011
Mechanism and cellular function of Bud6 as an actin nucleation-promoting factor.
November 1st, 2011
Determinants of endocytic membrane geometry, stability, and scission
March 8th, 2011
The formin DAD domain plays dual roles in autoinhibition and actin nucleation.
March 9th, 2010
A "primer"-based mechanism underlies branched actin filament network formation and motility.
March 15th, 2008
Stochastic severing of actin filaments by actin depolymerizing factor/cofilin controls the emergence of a steady dynamical regime.
April 1st, 2007
Attachment conditions control actin filament buckling and the production of forces.
March 18th, 2007