Regulation of Mical Redox Post-Translationally-Driven F-Actin Cytoskeletal Dynamics

The actin cytoskeleton is critical for multiple diverse cellular behaviors, including the ability of an axon to form, extend, navigate, and synapse with its target. Therefore, an important goal is to understand the mechanisms that regulate it. We have been studying one of the largest families of extracellular repulsive guidance cues, the Semaphorins, which were identified in part based on their ability to dramatically dismantle F-actin. More recently, we identified a new actin regulatory protein Mical, which directly associates with both the Semaphorin receptor Plexin and F-actin to post-translationally oxidize actin on its conserved methionine-44 and methionine-47 residues, inducing both F-actin disassembly and altered actin polymerization. Our work has also revealed that this Mical-mediated actin regulatory process is reversible by a specific methionine sulfoxide reductase enzyme called SelR/MsrB. Thus, we have identified an unusual new actin regulatory system - which I sought for my dissertation research to focus on better understanding. I now find that each human MICAL family member, hMICAL-1-3, similar to Drosophila Mical, directly induces F-actin dismantling and controls F-actin-mediated cellular remodeling. Thus, the MICALs are an important phylogenetically-conserved family of catalytically-acting F-actin disassembly factors. I also investigated how this new actin regulatory system fits with classically-studied actin regulatory proteins. Employing a simple biochemical screen, I identified two proteins - cofilin and tropomyosin - that modulate Mical-mediated F-actin disassembly. Further investigation revealed that Mical synergizes with cofilin to rapidly and efficiently dismantle F-actin in a redox regulated manner and that this synergism is also necessary and sufficient for F-actin disassembly in vivo - for remodeling cells, wiring the nervous system, and orchestrating Semaphorin/Plexin repulsion. In contrast, I find that tropomyosin - known to decorate F-actin within specific cellular compartments and at different developmental stages ¬- protects F-actin from Mical-mediated disassembly by stabilizing Mical-oxidized F-actin. Likewise, changing the levels of tropomyosin in vivo results in similar alterations to Mical-mediated F-actin/cellular remodeling suggesting a previously unknown mechanism controlling the plasticity of the actin cytoskeleton with important tissue-specific and developmental/age-related connotations. Thus, my findings provide new insights into the workings of this MICAL-mediated reversible Redox actin regulatory system including its importance to cell, developmental, and neural biology.