Research Group at Institut Jacques Monod
Summary of research:
The assembly of actin filaments is highly regulated in cells, giving rise to various well-controlled architectures (cortex, lamellipodia, filopodia....) and resulting in the specific deformations and movements that are required for many processes (cell motility, cell division, endocytosis, vesicle transport…). Actin-binding proteins are key regulators controlling how, where and when actin monomers will assemble into filaments. Understanding their molecular action can be very challenging, especially when multiple activities coexist within a single protein or within a stable complex. In addition, new regulatory effects can emerge from the combined action of several proteins. Our first aim is to decipher these biochemical regulatory activities, at the level of individual reactions.
Mechanics play a central role, as the actin cytoskeleton generates and transmits mechanical forces throughout cells and tissues. Growing evidence shows that the cytoskeleton also senses mechanical forces, and could be directly involved in the conversion of mechanical information into chemical signals. Yet very little is known about the elementary processes of actin mechano-sensitivity. Our goal is to understand how mechanical constraints affect actin dynamics and the action of regulatory proteins.
Actin filaments grow anchored by one end to the bottom of a microfluidic flow cell and align with the flow. Left: sketched perspective of the microchamber (top) and side view of the filaments (bottom). Right: epifluorescence microscope image.
In order to address these questions and unveil the elementary mechanical and chemical processes regulating actin assembly, single filament experiments are essential, in addition to standard biochemical assays. We have developed a microfluidics setup for the observation and manipulation of individual filaments, in combination with different optical microscopy techniques and optical traps. This technique enables us to observe actin filaments while changing their chemical environment, and to apply tensile forces up to a few picoNewtons to these filaments.
We are constantly improving this experimental microfluidics-based approach. So far, we have used it to shed light on issues like ATP hydrolysis in filaments, the action of profilin at the barbed end, the occurrence of pauses during filament depolymerization, the synergy between Spire and formin activities at the barbed end, and formin mechano-sensitivity.
