Katrín Möller, Juan Ouyang, Kaan Okay, Stefán Pétursson, Kasper D. Hansen and Hans Tómas Björnsson
Introduction: The epigenetic machinery (EM) includes over 300 proteins that regulate gene expression and chromatin accessibility. Mutations in EM genes commonly lead to intellectual disability in carriers, suggesting an important role in neurodevelopment. One hypothesis is that EM-components act as barriers, where timely placement or removal of individual EM-barriers allow cells to pass through different developmental stages. However, the roles of individual EM genes in this process are currently unclear. We aim to explore this hypothesis and identify shared transcriptional patterns underlying EM-related neurodevelopmental disorders.
Methods: We use a neurodevelopmental model where neuronal progenitor cells (NPCs) are differentiated into neurons in vitro. CRISPR/Cas9 is used to knock-out individual EM-genes in this model and both bulk and single-cell RNA sequencing (scRNAseq) are used to evaluate transcriptional changes. Immunofluorescence and western blotting are used to assess neurodevelopmental defects after individual EM knock-outs.
Results: RNA sequencing of our model revealed that many EM-genes are expressed in NPCs but are either up- or downregulated during neurodevelopment, supporting a barrier-like behavior. Furthermore, the disruption of many EM-genes causes abnormal neurodevelopment, including precocious differentiation. We are performing a scRNAseq experiment on our model, after a pooled knock-out of 60 disease-causing EM genes. This will reveal if there are shared transcriptional abnormalities amongst multiple EM knock-outs.
Conclusions: Our results revealed that many EM-genes show a barrier-like behavior during neurodevelopment and disrupting these genes causes premature differentiation of progenitors. By identifying shared transcriptional abnormalities after EM-disruption, our results could contribute to the development of treatments for EM-related disorders.
