Actin provides the critical driving force for muscle contraction and myoblast fusion. Turnover of actin structures and dynamic balance between actin arrays and cables has been implicated in muscle deterioration and regeneration. The central player in the formation of branched actin structures is Arp2/3, a protein complex containing two actin-related proteins that binds to the side of an existing actin filament and nucleates the formation of a new filament at a 70 degree angle. Force generation requires filament branching and elongation, but also regulated disassembly, leading to a treadmilling array. Key players in the disassembly reaction are cofilin, coronin, and Aip1, which disassemble ADP-containing actin filaments. However, the mechanism responsible for the removal of Arp2/3 and subsequent actin turnover remains mysterious. Cofilin has been directly associated with myopathies, and has been implicated in muscle regeneration processes. Recent investigation in the host lab has uncovered a role for GMF (glia maturation factor), a novel member of the cofilin superfamily, in removing Arp2/3-containing branch-points. This protein is structurally similar to cofilin, but does not bind actin, and instead binds Arp2/3 complex. Further, it is conserved from yeast to humans and is ubiquitously expressed, suggesting that it has a central and conserved role in regulating the actin cytoskeleton. Here, I propose to investigate the mechanism of GMF function and how its effects are coordinated with nucleotide hydrolysis on Arp2/3, previously shown to be important for regulating the timing of branch removal. I also propose to untangle a poorly understood set of interactions between GMF and coronin. Using TIRF microscopy, single molecule analysis, and an in vitro assay that reconstitutes actin-based motility, I will characterize the effects of GMF and other actin-turnover factors on the remodeling of arrays. The goal of these studies is to understand how the biochemical properties of an individual protein influence the behavior of a more complex actin-based system. The outcome of these studies will provide important new insights into the mechanism of actin turnover.
|Program type||Postdoctoral Fellowship|
|Effective start/end date||01/01/2011 → 12/31/2012|