The role of axonal mRNA translation in Amyotrophic Lateral Sclerosis (resubmission)

Amyotrophic lateral sclerosis (ALS) is a devastating disease caused by the degeneration of motor neurons leading to paralysis and death usually 2-5 years after onset of symptoms. Currently no cure or disease-modifying therapies exist, partly because we still lack basic understanding of the root-cause of the disease. To devise better therapeutic strategies, we need a greater understanding of the molecular and cellular mechanisms responsible for this selective neuronal degeneration. The vast majority of cases are sporadic while 5-10% are inherited. Studying the gene alterations that cause the disease in a rare few cases gives important clues into the causes. Functional studies of mutations that cause the disease have begun to implicate a number of mechanisms. One key mechanism that is emerging is the dysregulation of RNA biology. Current evidence indicates that multiple aspects of mRNA formation and handling are affected in disease conditions, including expression, maturation, stability, subcellular distribution and translation. The key question now is why disruption of mRNA handling leads specifically to the demise of motor nerves to cause ALS. A unique characteristic of motor nerves is their extraordinary length; many times longer than most other nerve types. In recent years, it has become clear that mRNAs can travel to the nerve extremities where they provide templates for local translation to support essential synaptic functions. Bringing these ideas together, we hypothesise that disruptions to the myriad of normal RNA regulatory processes leads to alterations in the population of mRNAs available for translation in motor nerve termini is an important trigger in how ALS starts. To explore this idea, we will determine which mRNA molecules are underdoing translation in adult motor nerve axons in vivo. We will then seek to understand how this profile changes during ageing and under disease conditions. Importantly, this work will go beyond the in vitro conditions of cultured cells by conducting our analysis in an animal model, Drosophila melanogaster. We will address these fundamental questions using the disease-relevant cell type (motor neurons) in their native environment, in an intact neuromuscular circuit. For time, cost and ethical considerations, invertebrate models provide many advantages yet have proven functional similarities to mammalian motor nerves. Moreover, using a genetically tractable animal model will allow us to define the pathological relevance of the mRNAs that are dysregulated by determining their functional impact on the disease-relevant circuits at an organismal level. This type of discovery research is essential to lay the foundations for a clearer understanding of the disease cause to develop more effective therapies.

Dysregulated RNA biology has emerged as a major driving force in the pathogenesis of Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia. While it is still unclear why motor nerves are particularly vulnerable in ALS, their extreme length further implicates axonal trafficking processes to be involved. The observed 'dying-back' pathology also supports that cellular disruption begins at the distal nerve terminus. Thus, we hypothesise that defects in processes such as transport and translation of certain mRNAs in motor neurons is an important contributor to neurodegeneration in ALS. In this proposal we aim to identify mRNA populations actively undergoing translation (translatome) specifically in the axons and synapses of motor nerves in vivo. We will use the model organism Drosophila to study the disease-relevant neuron type in situ, and to exploit its unparalleled genetic tractability. We have established a novel method to extract tagged ribosomes specifically from motor nerve axons. In this study, we will 1) determine the MN axonal translatome under normal conditions over the fly life-course, 2) investigate how axonal and somal translation is affected by TDP-43 mutations and C9orf72-related repeats during disease progression and 3) determine the relative contribution of different dysregulated axonal mRNAs to pathogenicity by assessing their functional impact on the disease-relevant circuits at an organismal level. The results of this project will provide new insights into how dysregulated RNA biology specifically impacts on motor neuron survival.

Both ALS and FTD-related disorders have an annual incidence rate of 1 to 2 per 100,000 people and approximately 10,000 individuals suffer from ALS or FTD at any time in the United Kingdom. A large proportion of patients with ALS onset develop FTD, and conversely, many patients with FTD will suffer later on from ALS symptoms. Whilst no treatment is available for FTD, the current standard of care for ALS patients, riluzole, only marginally extends survival. As advancing age is the main risk factor for ALS/FTD, the socio-economic impact is set to rise significantly in the future because of the aging population. In addition to care provided within the NHS, a significant amount of care for patients with ALS/FTD takes place in the community, often with support of charitable organisations such as the Motor Neurone Disease Association and Dementia UK. Thus, ALS/FTD places a significant public and private socio-economic burden on the UK.

In the short term, the main beneficiaries of knowledge arising from this research will be the people employed on this grant, and academic and clinical neuroscientists, particularly those with an interest in ALS, other neurodegenerative diseases and dementia such as FTD. The employment and training of research staff involved in the project will benefit their careers both by enhancing specific research skills and experience, as well as a range of transferable skills such as analysis and problem solving, interpersonal and leadership skills, project management and organisation, information management, self-management, and written and oral communication.

In the medium to long term we anticipate that our research will have a wider impact and benefit the pharmaceutical industry, neurodegenerative disease sufferers and their families, and society as a whole. Our research seeks to identify novel therapeutic targets in ALS (and other neurodegenerative diseases and dementia) that can be developed into better disease-modifying therapies. The effect of better therapies will be most tangible for patients and their immediate families. Improving the symptoms and halting or slowing the progression of disease will have an enormous impact on their quality of life. Such a breakthrough would free up considerable resources in the NHS that are currently allocated to ALS/FTD and are set to rise in the future.

The University of Cambridge and MRC could benefit from licensing of any patents granted based on our findings, enhanced REF performance and increased reputation (via publications, press releases, international presentations etc.).

In addition to its impact on health and wellbeing, the public, and the public sector our research may also have an economic impact via the commercialisation of our results (e.g. therapies based on mitochondrial calcium modulating drugs) and/or spinout companies. Accordingly, the main commercial beneficiaries would be the pharmaceutical industry with whom we will engage in the later stages of drug development.

Finally, as the outcome of this research may also be applicable to other neurodegenerative disease such as Alzheimer's, Huntington's and Parkinson's disease, the potential impact may be much higher.