Autoinhibition and Activation Mechanisms for MICAL Monooxygenases in F-actin Disassembly

Abstract

MICAL (Molecule Interacting with CasL) proteins represent a unique family of actin regulators crucial for synapse development, membrane trafficking, and cytokinesis. Unlike classical actin regulators, MICAL proteins possess enzymatic activity, catalyzing the oxidation of specific residues within actin filaments to induce robust filament severing. The potent activity of MICAL proteins requires tight control to prevent extensive damage to the actin cytoskeleton. Due to limited structural information on full-length MICAL proteins, the molecular mechanisms governing MICAL autoinhibition and activation remain elusive. Here, we reported the cryo-EM structure of full-length MICAL1, unveiling a head-to-tail interaction that allosterically blocks enzymatic activity through the binding of the C-terminal Rab-binding domain (RBD) to the N-terminal monooxygenase domain. The structure also reveals the assembly of C-terminal domains via a tripartite interdomain interaction, stabilizing the inhibitory conformation of the RBD. Our structural, biochemical, and cellular analyses elucidate the transition from the autoinhibited to activated conformation of MICAL1 in response to Rab8-subfamily GTPase. Dual Rab-binding to the RBD induces the conformation rearrangement of the RBD and synergistically promotes the autoinhibition-to-activation transition, revealing intricate activity regulation mechanisms for MICAL proteins. Furthermore, our mutagenesis study of MICAL3 suggests a conserved autoinhibition and activation mechanism among MICAL proteins.