Investigating chromatin misfolding as a pathogenic mechanism in neurodevelopmental disorders.

The brain is a complicated organ that requires careful organisation when it first develops in embryos. Small mistakes, such as not making a normal balance of neurons (electrical cells of the brain), can lead to common and severe disorders. One example of this is epilepsy, which affects ~1% of the UK population. Unfortunately, it is linked >1100 deaths every year and costs the NHS ~£1 billion. Even more worrying is that around a third of epilepsy patients have drug resistant forms, which leaves these people with risky treatments, such as brain surgery. To improve the care of these people, we need new treatments or tests to predict who will develop severe forms. By studying how epilepsy brains develop (and comparing them to normal ones) we will be able to identify new drug and test targets.

Research has already given us clues about how epilepsy brains may develop differently. Normally, when our brain forms, neurons multiply and specialise - some stimulate electrical activity, others inhibit it. The intricate patterns of electrically stimulatory and inhibitory neurons are important for co-ordinated electrical activity in the brain. In epilepsy, however, it is thought that there may be an excess number of stimulatory neurons, likely because there is a reduced number of inhibitory ones forming. This creates difficulties in co-ordinating the electrical activity of the brain and can lead to overactive, uncoordinated brain activity and seizures.

We are starting to uncover the mechanisms for why there is a reduced number of inhibitory neurons. Scientists have identified a list of genes that when mutated give rise to syndromes in which drug-resistent epilepsy is particularly common. Interestingly, many of these genes help to control how DNA is folded and stored, which determines the genes that make get to make proteins and thus instructions given to the cell. The main instruction - telling the neuron whether to become electrically stimulatory or inhibitory - gets very confusing when DNA is not stored correctly. We believe that DNA-folding genes are mutated that far fewer neurons get the instruction to turn into electrically inhibitory neurons.

To investigate our ideas, we propose to study two DNA-folding genes (called FOXG1 and CHD2) that when mutated give rise to syndromes in which drug-resistant epilepsy is very common. Using stem cells that can be guided to make any cell type in the adult body, we will genetically modify them so the DNA folding genes FOXG1 and CHD2 are mutated. We will then grow these stem cells as 3D collections that resemble parts of the developing human brain where inhibitory neurons are made (a new technology known as brain organoids). This technique will allow us to study human brain development as accurately as is currently possible and without the need to use animals. Using brain organoids and stem cells, we will then look at how DNA is folded in neurons where there is no FOXG1 or CHD2. If we see abnormal DNA folding patterns, we will then genetically modify another group of stem cells to try and recreate the abnormal DNA folding patterns (but with normal FOXG1 and CHD2 protein). If this new cell line creates brain organoids with low numbers of inhibitory neurons, then this will be very strong evidence that abnormal DNA folding is a way in which epilepsy and neurodevelopmental disorders generally can develop.

The findings of this study could have strong scientific and medical implications. We will provide some much-needed insight into how genes with DNA folding functions control how our brains develop. Similarly, it will be important medically as it opens up the possibility that this is a mechanism for how a wide range of other conditions (associated with abnormal brain development in embryos) develop. This will in turn provide numerous targets for future research looking to find new targets for diagnostic tests or drugs for novel treatments.

Atypical neurodevelopment gives rise to a range of prevalent and burdensome disorders. Epilepsy, for example, affects ~1% of the UK population. Improving our understanding of the pathogenesis of neurodevelopmental disorders will identify new treatment and diagnostic targets for these disorders, which is important for epilepsy where 36.3% of patients experience drug resistance and poor outcomes.

How can we improve our understanding of the pathogenesis of neurodevelopmental disorders? One way is to study the function of genes that, when mutated, give risk to neurodevelopmental conditions, such as FOXG1 (transcription factor) and CHD2 (part of a chromatin remodeller complex). Both control neural progenitor cell (NPC) differentiation and studies indicate they control chromatin folding (the 3D way DNA is stored). Interestingly, epilepsy is extremely common in syndromes arising from FOXG1/CHD2 mutations, and abnormal neuron differentiation patterns (excitatory-inhibitory imbalance) is implicated as core to its pathogenesis.
A detailed molecular mechanism linking atypical NPC differentiation to FOXG1/CHD2 mutations remains unknown.

Here, I hypothesise loss of CHD2 or FOXG1 causes chromatin misfolding which drives imbalanced NPC differentiation, ultimately giving rise to excitatory-inhibitory imbalance

To investigate my hypothesis, I have 3 aims:
- 1. determine how FOXG1-/CHD2-loss influence transcription in NPC subpopulations
- 2. determine how FOXG1-/CHD2-loss alter NPC chromatin folding patterns
- 3. identify FOXG1-/CHD2-regulated chromatin folding patterns controlling NPC transcription and differentiation

This exciting project offers numerous academic and medical opportunities. It will help to uncover a possible molecular mechanism behind epilepsy and neurodevelopmental disorders. This is essential to improving the care of people with disorders like drug-resistant epilepsy, by identifying new treatment and diagnostic targets for future research