L’équipe Romet-Lemonne/Jégou has just published a new article in Biophysical Journal :
(ABSTRACT VOLUME 122, ISSUE 3, SUPPLEMENT 1, 166A-167A, FEBRUARY 10, 2023)
Actin filaments are one of the indispensable components of almost all cell types. These semi-flexible filaments constitute an important part of the cytoskeleton and give the cell its mechanical properties. Among various structures that actin filaments form, branched actin network plays a crucial role in many cellular processes such as cell motility and endocytosis. Branched actin filaments are formed when an activated Arp2/3 complex binds to the side of an existing filament (mother) and generates a new filament branching off the mother. These filaments are subjected to force when they are pushing against the membrane or as a result of myosin motors pulling on them. Furthermore, biochemical conditions, namely actin-binding proteins, can regulate the dynamics of branched actin networks, notably the stability of branch junctions. Here we use a microfluidics approach to reconstitute individual branched actin filaments from purified proteins in-vitro. Thanks to microfluidics, we can grow filaments branching off from the side of the mother filaments and pull on them with different orientations, varying continuously between 0 and 180 degrees. Such an approach allows us to characterize the effect of force orientation, as well as force magnitude, on debranching in the presence of different regulatory proteins. Furthermore, we are able to follow the fate of Arp2/3 after debranching. Contrary to previous observations, we find specific physiological conditions in which Arp2/3 can remain attached to the mother filament after branch departure. Unexpectedly, in this type of debranching, the remaining Arp2/3 complex is able to nucleate a new branch and contribute to the turnover of the network.