Regulation and resilience of the neuronal microtubule cytoskeleton in health and disease

Our brains are built from billions of specialised cells called neurons. The many complex tasks that our brains perform, including thought and memory, occur because neurons make connections with each other that allow them to communicate. Early in brain development, immature neurons are not connected to each other and must navigate to exactly the right position to correctly integrate into the brain's communication network. Healthy brain function throughout our lives depends on the connections between our neurons being well maintained. Severe human diseases can occur if neuron connectivity and operation breaks down at any stage. Inaccurate neuron movement during brain development can cause intellectual disability, epilepsy and early death. Incomplete maintenance of neuronal function as our brains mature into adulthood can also cause neuropsychiatric illnesses including schizophrenia. Breakdown of neuronal function as we age can cause neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS). In all these disease scenarios, there remains much to learn, and work in my lab is seeking to understand the machinery that supports neuronal health during development and as we mature.

In the same way as our body has a skeleton that provides us with support and strength, neurons have a skeleton - called the cytoskeleton - which also gives them support and strength. The cytoskeleton is involved in many important aspects of neuronal life, and is part of the machinery that drives neuron movement during development, along with maintenance of connectivity and communication in mature neurons. Breakdown or disruption of the neuronal cytoskeleton is associated with developmental syndromes, neurodegenerative diseases and neuropsychiatric illnesses. Studying the cytoskeleton machinery is important so we can understand both how healthy neurons operate and how machinery malfunction causes disease.

This project will focus on a part of the cytoskeleton called microtubules. These are long cylindrical structures that act like scaffolding inside the neuron and also act as tracks along which molecular transport motors carry cargo within the neuron. The organisation and stability of the microtubule machinery, together with the particular type of cargo that is carried along it, defines how the neuron functions. We would like to understand how the neuronal microtubules are assembled and maintained to help neurons undertake their many complex tasks within the brain. My research team studies the three-dimensional structure of microtubules, because knowing what they look like can help us understand how they work. We use a very powerful microscope called an electron microscope to take pictures of individual microtubules that have either been assembled in a test tube or form within a living neuron. We then use computers to combine these electron microscope pictures to calculate the microtubules' three-dimensional shape. By using information from patients with diseases that disrupt the microtubule machinery, we will be able to map disease-causing defects to particular machinery components.

In the future, knowledge arising from our work may allow us to target and repair the broken parts of the cytoskeleton machinery in diseased or damaged neurons. Such understanding could also shed light on new treatments for dementia, stroke and physical injury.

The operation of the human brain depends on connectivity between the billions of neurons from which it is built, which in turn depends on precise localisation of neurons during development. The microtubule (MT) cytoskeleton plays critical roles in diverse neuronal activities, and its importance in human health is spotlighted by the neurological diseases that are caused or influenced by MT disruption. Studying the molecular operation and regulation of MTs is thus essential to understand healthy neuron development and operation, to dissect the consequences of its disruption in diseases and for development of brain repair technologies.

This research programme will uncover the molecular mechanisms of essential neuronal cytoskeleton components that are disrupted in disease and elucidate the effects of this disruption on neuronal ultrastructural organisation and function. We will investigate three families of MT regulators - DCX-MAPs, CAMSAPs, MAP7s - to reveal how they control MTs and influence trafficking through interactions with motors. Using reconstitution of individual components, the ways in which MTs are initiated, stabilised and their integrity maintained will be explored. Cryo-electron microscopy will be used to visualise the interactions that mediate these activities at high resolution. We will also use cryo-electron tomography (cryo-ET) to determine unique MT structures, including the response of binding partners to structural defects, and to MT-binding drugs. Dynamic interactions of regulators with MTs will be probed using biophysical methods and hypotheses will be tested using protein engineering. We will use cryo-ET to study MTs in situ in neurons, will elucidate the impact of disruption of specific MT regulators on their ultrastructure and organisation, and will visualise the response of neuronal MTs to MT-modulating drugs . We will thereby uncover regulatory mechanisms of MT resilience with implications for human health.