The Glycine Cleavage System in Brain Development, Function and Disease

We aim to understand how impaired function of the glycine decarboxylase (GLDC) protein leads to disorders of brain development and function. GLDC acts to break down a small molecule called glycine, allowing part of the molecule to enter a network of chemical reactions known as folate metabolism, which is needed in almost all cells for many different functions. Loss of GLDC activity, resulting for example from an inherited genetic mutation, leads to accumulation of excess glycine and to suppression of folate metabolism. Our goal is to understand how these changes lead to life-threatening brain diseases that arise both before and after birth. These disorders include common birth defects such as neural tube defects (NTDs) and congenital hydrocephalus, as well as the severe childhood disease Non-Ketotic Hyperglycinemia (NKH). Abnormal folate metabolism contributes to NTDs, hydrocephalus and NKH. Understanding causal links is also of broader relevance as folate metabolism is implicated in a range of other disorders (e.g. birth defects, cancers, and neurological disease), and may be altered by inherited genetic changes and other factors such as diet.

NTDs occur in approximately 1-2 per 1,000 pregnancies, due to incomplete formation of the neural tube, which later develops into the brain and spinal cord. The brain and/or spinal cord of the fetus become irreversibly damaged, resulting in death around birth or long-term disability in surviving children. Because of the many possible contributory factors, the exact cause of NTDs in each individual is usually unknown. Working out how GLDC defects cause NTDs provides an opportunity to gain a better understanding of the link between folate metabolism and NTDs.
Congenital hydrocephalus affects 0.5-1 per 1,000 babies and can lead to brain injury owing to increased pressure of cerebrospinal fluid within the brain. Hydrocephalus caused by GLDC mutation results from a blockage of fluid flow due to incorrect building of the fetal brain structure. Work leading to the current study shows that this is a result of impaired folate metabolism. The next step is to understand exactly when and in which cells the process of brain development goes wrong, which of the outputs of folate metabolism are disrupted, and whether these changes are responsible for hydrocephalus. In addition to contributing to NTDs and hydrocephalus, GLDC mutation is the major cause of NKH, an inherited disease that affects around 1 per 50,000 babies and causes epilepsy, profound development delay and early death. The relative contribution of excess glycine and impaired folate metabolism to different aspects of NKH is not well understood and we will address this question.

The risk of NTDs can be substantially reduced if the mother takes supplemental folic acid (related to folate) supplements before or during early pregnancy. However, a substantial number of NTDs are resistant to folic acid and there is a need to identify additional therapies. A key aim is to identify new preventive therapies for NTDs and hydrocephalus which may be used individually or in combination with folic acid. In families where genetic risk factors have been identified this also means that family-specific therapies may be offered. Current treatments for NKH are not effective and there is no cure. In order to implement new therapies it is important to understand whether some features of NKH result from abnormalities that already manifest before birth and whether these can be corrected.

We will address key outstanding questions about the mechanisms underlying NTDs, hydrocephalus and NKH using mouse and human cell models lacking function of GLDC. These models provide an opportunity to determine the precise effects of GLDC defects on folate metabolism and linked biochemical reactions, how these lead to changes in precursor cells that are essential for brain development and whether particular groups of cells are abnormal in the brain after birth.

This programme aims to understand how suppression of folate one-carbon metabolism (FOCM) leads to abnormal brain development and function. We focus on glycine decarboxylase (GLDC), a component of FOCM which has been implicated in neural tube defects (NTDs), hydrocephalus and Non-Ketotic Hyperglycinemia (NKH). Using genetic models in mouse and human iPSC models our main goals are:
1. To determine the effect of GLDC loss of function on metabolism, cellular phenotypes and patterning in embryonic neuroepithelium and neuronal precursor cells. We will use: (i) functional assays of energy metabolism, metabolic tracing and imaging of fluorescent reporters; (ii) assays of cell cycle and redox balance; (iii) evaluation of one-carbon donors and other metabolic intermediates for prevention/exacerbation of NTDs; (iv) gene expression analysis.
2. To test the requirement for GLDC in prevention of structural malformations that lead to hydrocephalus. Morphometric and gene expression analysis of diencephalon will determine where/when defects first arise, with conditional genetic rescue to localise the need for region-specific GLDC function. Potential causative metabolic and cellular defects will be examined by testing preventive methods.
3. To identify the role of GLDC in postnatal brain development. Cell cycle analysis in pre-natal progenitors and immunostaining in juvenile brain will reveal cortical layering anomalies. The requirement for GLDC in post-natal NPCs will be determined by lineage tracing, immunohistochemistry and single cell transcriptomics. Co-cultures will reveal whether GLDC loss in astrocytes has deleterious effects on metabolism or redox balance in adjacent neurons.
Identification of pathogenic mechanisms underlying GLDC-related disorders will provide an opportunity for rational design of potential therapeutic approaches which will be evaluated for prevention of NTDs and hydrocephalus, and for amelioration of cellular and metabolic defects associated with NKH.