Title: Investigating mechanisms of RNA dysregulation in C9orf72-associated motor neuron disease and frontotemporal dementia
Funder: The Academy of Medical Sciences

Abstract: Background: Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are fatal disorders characterised by neuronal degeneration. ALS affects 1 in 300 people in the UK and FTD is a common cause of young-onset dementia, but there are no curative treatments. A hexanucleotide expansion in C9orf72 is the commonest genetic cause of ALS and FTD. Previous work in preclinical models suggests that neurotoxicity is mediated by dipeptide repeat proteins (DPRs) generated by repeat-associated non-ATG translation. My preliminary evidence shows that DPRs bind specific RNA molecules in human cells, indicating a novel disease mechanism. I hypothesise that this mechanism will lead to neurotoxicity. Investigating its impact will lead to novel insights. I have found DPRs bind to nucleolin RNA very frequently. Nucleolin is an RNA-binding protein and recent evidence demonstrated its novel role in regulating neuronal size and mRNA translation in neurites, which is an important potential mechanism in ALS. Aims: Using induced pluripotent stem cell (iPSC)-neurons, I will evaluate DPR-RNA interactions, effects on transcription, and how nucleolin is affected. Methods: I will express DPRs in iPSC-neurons. Using genome-wide individual nucleotide resolution UV-crosslinking and immunoprecipitation (iCLIP) alongside RNA sequencing, I will investigate specific binding of DPRs to RNA and the effect on the transcriptome. I will validate hits using qRT-PCR, western blotting and immunofluorescence. I will investigate subcellular localisation of nucleolin and neurite length. Implications: These experiments will provide mechanistic insights into a common form of ALS/FTD and deliver data on a novel disease process, with potential for translation to new therapies.

Lay abstract: Motor neuron disease (MND), also known as amyotrophic lateral sclerosis (ALS), is a devastating, fatal condition. One in 300 people in the UK are affected by this disease. Frontotemporal dementia (FTD) is a common cause of young-onset dementia. There are no effective treatments for these diseases. A gene called C9orf72 causes a significant proportion of MND/ALS and FTD. Mutated C9orf72 leads to abnormal proteins being made which are lethal and lead to cell death. In my initial work in human cells, I made the exciting new discovery that these abnormal proteins bind to RNA, which are messenger substances that allow the genetic code, DNA, to be made into proteins. It is critical to understand the impact of this binding between the abnormal proteins and RNA in nerve cells, as this will increase our understanding of these diseases. I have also found that the abnormal proteins frequently bind to the RNA of a gene called nucleolin. Nucleolin is an important protein that has many roles, including in changing the size of nerve cells. However, we do not fully understand how nucleolin is affected in C9orf72 nerve cells. I will study if these abnormal proteins lead to harmful effects on RNA in neurons made from human stem cells, and how nucleolin is affected. Understanding this will help us discover why these disease happens, and potentially find new ways to treat them. I hope this research will allow us to find treatments which could improve the lives of people with MND/ALS and FTD.

Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: NEUROLOGICAL