RNA metabolism in pathology of AOA2/ALS4 neurodegenerative disorders

All living organisms contain genetic information in a form of genes, which code for proteins - key building blocks of life. During the process of gene expression information from the gene, encoded in a DNA sequence is transferred into messenger RNA (transcription) followed by removal of non-coding intervening sequences (splicing), and decoding of this molecule into protein (translation). Each gene expression step is subjected to multiple regulatory events ensuring that the final protein product is correct. Now it became clear that key regulatory elements in gene expression are represented by the RNA. In this research project we will investigate the mechanism underlying the pathology of ataxia oculomotor apraxia type 2 (AOA2) and amyotrophic lateral sclerosis type 4 (ALS4), highly disabling inherited disorders, characterized by degeneration in the brain and spinal cord, causing progressive muscle weakness and finally atrophy. The gene mutated in these diseases encodes a protein called senataxin. Since the discovery of AOA2/ALS4 mutations in 2004, little progress has been made to characterize the function of senataxin protein or molecular mechanisms of AOA2/ALS4 pathology.
To study the function of senataxin protein in human cells, we will combine our expertise in RNA field with cutting edge high throughput technology in gene expression. In particular, we will perform experiments in human cells, which lack senataxin or were derived from patients' tissues. We hypothesise that AOA2/ALS4 pathologies arise due to defects in senataxin function associated with resolving connections between RNA and DNA, appearing during the process of transcription. If RNA/DNA molecules are not resolved properly, their accumulation can be detrimental to the cells causing pathologies. Initially, we will characterise the defects in transcription and RNA processing observed in cells with senataxin mutations using modern technologies analysing whole cell genome. We will also determine if RNA/DNA profile is perturbed in AOA2/ALS4 patients, and how it contributes to neurodegeneration. Furthermore, we will investigate the role of senataxin in transcription and RNA splicing. We will also identify new senataxin-interacting partners and study their contribution towards gene expression process and AOA2/ALS4 pathology.
Our project focuses on the function of senataxin, providing molecular clues for the pathology of AOA2/ALS4 diseases. It's becoming apparent that RNA processing may represent a common pathogenic mechanism involved in many neurodegenerative disorders. Around 50% of all characterised human genetic diseases are associated with inappropriate transcription/RNA processing. This emphasises the importance of fundamental molecular biology approach of our research towards understanding AOA2/ALS4 pathology. Therefore, the development of novel RNA-based therapies, improving the health and quality of life of AOA2/ALS4 and other neurodegenerative patients with defects of RNA processing, is a likely prospect of our research.

Ataxia oculomotor apraxia type 2 (AOA2) and amyotrophic lateral sclerosis type 4 (ALS4), are highly disabling disorders, characterised by neurodegeneration in the brain and spinal cord, causing progressive muscle weakness and finally atrophy, arising due to mutations in human senataxin protein. Senataxin shares homology with the yeast RNA/DNA helicase Sen1p and it has been suggested that senataxin may be involved in the regulation of gene expression. This research project will uncover molecular function of human senataxin protein using a combination of our expertise in RNA field and the cutting edge high throughput technology. In the first part of this project we will study transcriptional and RNA processing defects associated with AOA2/ALS4 mutations genome-wide. We propose that the function of senataxin in human cells is related to its RNA/DNA helicase function and its ability to unwind RNA/DNA hybrids (or R-loops) formed between nascent RNA transcripts and single stranded DNA template behind the elongating Pol II. We will investigate the effect of senataxin on genome-wide distribution of R-loops and study their contribution to the neurodegenerative phenotype, employing Solexa sequencing following DNA-immuno-precipitation (DIP-seq). Secondly, we will define the molecular functions of R-loops and senataxin in splicing regulation. Finally,we will identify novel RNA and protein senataxin-interacting partners, using mass-spectrometry and iCLIP approach, followed by the study of their function in gene expression and AOA2/ALS4 pathology. The long term goal of our work is to create a foundation for the collaboration between basic and clinical science beneficial to therapeutic advancement of AOA2/ALS4 diseases and development of novel molecular therapies, improving the health and quality of patients' life.

The outcome of this research project will be beneficial to all AOA2/ALS4 patients affected by mutations in senataxin protein and also to neurologists and clinical geneticists, directly involved in treating these patients. Since the discovery of senataxin mutations in 2004, little is known about genotype/phenotype correlation in AOA2/ALS4 patients, making the correct diagnosis of these diseases in hospital practice quite laborious and difficult. Our work will make a major contribution to this knowledge, making the strategy for diagnosis of these diseases more efficient.
Development of any therapeutic treatments for AOA2/ALS4 patients bas been hampered by the lack of understanding of molecular role of senataxin protein in humans. This project will investigate the function of senataxin in human cells and therefore shed the light on the underlying pathology of AOA2 disease, which may ultimately provide new targets for therapeutic treatment. In the long term, we are hoping to have an impact on treatment practices of these diseases.
Proposed research project will be instrumental in the initiation of public awareness of human genetic disease research and corresponding therapeutic approaches. In particular, during the NIRG, we intend to communicate about the AOA2/ALS4 diseases to patients and healthy individuals through our research publications, scientific presentations and outreach work with the help of University of Oxford and Ataxia UK, Motor Neuron Disease Association and National Ataxia Foundation charities. The results of this research project are likely to have a big impact on the national health, thus enhancing quality of life and health of patients.