Genetic mutation in OXR1 gene linked to brain development issues

By Vijay Kumar MalesuNov 27 2023Reviewed by Benedette Cuffari, M.Sc.

In a recent study published in Genome Biology, researchers investigate the impact of a novel loss-of-function mutation in the oxidation resistance 1 (OXR1) gene on cellular functions, neurodevelopment, and molecular mechanisms in the human brain.

Study: A loss-of-function mutation in human Oxidation Resistance 1 disrupts the spatial–temporal regulation of histone arginine methylation in neurodevelopment. Image Credit: Kittyfly / Shutterstock.com

Background 

OXR1 is a conserved gene crucial for oxidative stress resistance, thus playing a vital role in various cellular processes and diseases. Although it is not a direct source of DNA damage, OXR1 indirectly affects DNA damage response systems, antioxidant processes, and neuronal protection.

Despite the established roles of OXR1 in a wide range of biological processes and its association with various diseases, the detailed molecular mechanisms, especially the structural and functional roles of its specific domains in human neurological conditions, remain largely unexplored and poorly understood.

About the study 

The current study involved in vitro studies using immortalized lymphoblast cell lines obtained from a human patient, a control line infected with Epstein-Barr virus, fibroblast cells, and osteosarcoma U2OS cells.

Cell viability and apoptosis were explored through assays involving hydrogen peroxide (H₂O₂) exposure, flow cytometry, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. The researchers also measured reactive oxygen species (ROS) and performed high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC–MS/MS) analysis of 8-oxo dihydroquinine (dG) in genomic DNA to assess oxidative stress.

Human induced pluripotent stem cells (hiPSCs) were generated from patient fibroblasts and differentiated into neural cells. Formulation of cerebral organoids and region-specific brain organoids presented an approach for studying the effect of the OXR1 mutations on brain development. This was confirmed by immunofluorescent staining, immunolabeling analysis, ribonucleic acid (RNA) sequencing, and proximity ligation assays that focused on the interaction between OXR1 and other proteins.

Study findings 

In the present study exploring the implications of OXR1 deficiency, three sisters born to first cousins exhibited developmental delays, hypoactivity, cognitive issues, and epilepsy from an early age. Generalized hypotonia, hyporeflexia, ataxia, and non-communicative conduct were reported during their clinical examinations. Furthermore, electroencephalograms demonstrated severe epileptiform activity, whereas magnetic resonance imaging (MRI) scans revealed cerebellar and corpus callosum degeneration. 

A homozygous variant found on exome sequencing within the OXR1 gene was absent in broader population DNA databases. The mutation excluded exon 18 within OXR1, which is required for supporting protein stability, especially its terminus C containing TBC/LysM-associated domain containing (TLDc) domain.

To understand the functional consequences of OXR1 deficiency, lymphoblast cell lines (LB) from the patient and a healthy sibling were studied. The patient cells exhibited diminished proliferation, heightened sensitivity to oxidative stress, and increased apoptosis.

Elevated levels of 8-oxoguanine in patient DNA indicated enhanced oxidative DNA damage. Furthermore, these cells exhibited increased mitochondrial DNA mutation frequencies and altered expression of stress response genes, thus highlighting the role of OXR1 in cellular growth, survival, and oxidative stress response.

The researchers also generated iPSCs from patient fibroblasts. These iPSCs, which lacked OXR1, showed significant deficits in neural differentiation, with irregular neural aggregate morphology, reduced neurite growth, and decreased expression of neuronal markers.

Transcriptome analysis during neuronal differentiation revealed that OXR1 influences gene expression, with a notable impact on genes associated with cerebellar atrophy, autism spectrum disorder, and schizophrenia. Pathway analyses indicated disrupted neural development pathways in OXR1-deficient cells.

OXR1 was also found to interact with protein arginine methyltransferases (PRMTs), which are key regulators of gene expression. OXR1 deficiency resulted in impaired histone arginine and lysine methylations, which are crucial for neurogenesis. The study findings also demonstrated that the OXR1 TLDc domain is vital for stimulating PRMT5-catalyzed histone methylation, thus underscoring its role in transcriptional regulation during neurogenesis.

To mimic OXR1 deficiency in a more complex model, human brain organoids derived from patient iPSCs were developed. These organoids exhibited structural and developmental abnormalities, including delayed neuronal layering and reduced formation of specific brain regions. OXR1 deficiency was also found to disrupt various histone modifications, thereby affecting early brain development, particularly in cortical and midbrain regions.