Title: Modelling regulatory mechanisms of RNP condensation in neuronal development: insights into neurodegeneration
Funder: The Academy of Medical Sciences

Abstract: The structure of RNA molecules is crucial in mediating post-transcriptional regulation through an interplay with RNA binding proteins (RBPs) that forms ribonucleoprotein complexes (RNPs). Long stretches of RNA can form scaffolds upon which RNPs can condense creating microscopically visible granules. In neurodegenerative conditions, (e.g. motor neuron disease), these condensates can become aberrant, although the mechanisms are largely unknown. This proposal aims to understand the mechanisms by which RNA structures and RBPs can form condensates, describe the effects on RNA function and ultimately build a predictive computational model. My hypothesis is that secondary structures in 3’ untranslated regions control condensation and this regulates neuronal mRNA localisation and stability. I will use neurons derived from mouse embryonic stem cells as a model system and Staufen, an RBP that binds double-stranded RNA, as a window into RNPs. First, to identify RNA secondary structures important for condensation, I will generate a transcriptome-wide atlas of structures bound by Staufen using hiCLIP. Then to deconvolve structural heterogeneity and understand the mechanisms at single molecule resolution, we will develop a new structure probing method using direct Nanopore sequencing. Second, to understand the functional implications of RNP condensation, I will use publicly available RNA localisation and stability data, together with new single molecule stability experiments to generate a catalogue of effects. Finally, I will use machine learning methods to integrate the data from the first two aims to derive a predictive model of RNP condensation. This will enable us to test pathogenic and therapeutic manipulations in silico.

Lay abstract: In all cells, genetic information encoded in DNA is transcribed into RNA, which forms the template for protein production. Controlling this process correctly is crucial for cells to work properly. This is particularly important in nerves, which have long extensions called axons. Quick responses are needed at the end of the axon when the nerve fires. To enable this, a pool of the relevant RNA molecules is kept there in standby ready to help generate proteins rapidly. Regulatory proteins bound to the RNA control this local pool. In people who suffer from diseases where nerves die, such as motor neuron disease, this mechanism becomes defective. Although we know some of the genes that are involved, we do not yet know how these defects cause the disease. I will study how the binding of proteins to specific structures in RNA controls the movement and the breakdown of RNA. First I will use a method we developed to detect these structures in mouse nerve cells grown in the lab. Then we will develop a new technology that can identify these in individual RNA molecules. This will help us understand the diversity of structures, how quickly they degrade and where they are located. Finally, I will use artificial intelligence methods to build a model that can predict the effect of binding to structures on the stability and localisation of RNAs. This will enable me to test the effects of disease mutations in the computer model and also work out how to fix them.

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