Structural and mechanistic basis of MICAL regulation

Descripción del proyecto

Perspectiva mecanística de la regulación de la señalización MICAL

Las enzimas MICAL son fundamentales para diversos procesos basados en el citoesqueleto, incluida la guía de axones, la morfología celular, la sinaptogénesis y la plasticidad neuronal. Se fijan directamente a filamentos de actina y participan en su desconexión dinámica. El proyecto financiado con fondos europeos MICAL tiene por objeto comprender los mecanismos moleculares subyacentes a la señalización MICAL, con un especial énfasis sobre su activación y autoinhibición. Los científicos utilizarán tecnologías de vanguardia para realizar una determinación a alta resolución de la estructura de las enzimas MICAL así como de otras proteínas, lo que permitirá obtener información importante sobre sus interacciones moleculares. Los resultados del proyecto abrirán nuevas posibilidades sobre la manipulación de la señalización basada en MICAL y ayudará a desarrollar tratamientos nuevos para los trastornos neurológicos.

Objetivo

MICALs are a family of redox enzymes that directly bind and disassemble actin filaments. An increasing body of evidence in recent years indicates that MICALs play essential roles in many tissues for myriad activities requiring discrete changes in the cytoskeleton, including axon guidance, cell morphology, synaptogenesis and neuronal plasticity. MICALs are a component of classical signalling pathways and are best known as effectors for semaphorin-plexin signalling. Although the field has made enormous advances in understanding MICAL function at the level of genetic and cellular experiments, our knowledge of the molecular-level mechanisms of MICAL signalling in cytoskeletal dynamics remains poorly understood.
The overarching aim of this proposal is to understand the molecular mechanisms underlying MICAL signalling. I aim to address two fundamental and long-standing questions: (1) How do MICAL proteins precisely turn their activity on and off? (2) How do cytoplasmic segments of plexins pass the signal on to MICALs and what mechanisms control signalling? I will use a hybrid approach integrating single particle cryoEM with high-resolution X-ray crystallography to determine the overall architecture of MICAL and to elucidate molecular mechanisms of MICAL autoinhibition. In parallel, I will aim to determine the high-resolution structure of MICAL in complex with plexin to dissect molecular mechanisms of MICAL activation.
All previous attempts to tease out the mechanisms governing MICAL signalling failed because full-length MICAL is expressed at a relatively low level in many expression systems. The project will greatly benefit from my previously developed protocol describing the production of MICALs in milligram quantities. Our findings will reveal fundamental principles of MICAL signalling, which will open new possibilities in the manipulation of cell signalling, and pave the way towards the treatment of MICAL associated neurological disorders.