Small molecule inhibitors of G9a reactivate the maternal PWS genes in Prader-Willi-Syndrome patient derived neural stem cells and differentiated neurons

Abstract

Abstract/SummaryPatients with Prader-Willi-Syndrome (PWS) display intellectual impairment, hyperphagia, and various behavioral problems during childhood that converge on a neurologic deficit. The majority of PWS patients have genetic deletions of the paternal 15q11–q13 chromosomal region, with their maternal PWS locus intact but epigenetically silenced by hypermethylation and repressive histone modulation of the PWS imprinting center (PWS-IC). Inhibition of the euchromatin histone methyltransferase G9a by small molecules has been recently reported to reactivate PWS genes in patient fibroblasts and a mouse model. However, it is unknown if inhibition of G9a could have similar effect in human PWS neural cells, the cell types that have direct pathophysiological relevance to PWS. Here, we use neural progenitor cells (NPCs) and cortical excitatory neurons derived from a patient iPSC to model PWS, and quantitatively profile the expression of PWS genes using a NanoString panel. We demonstrated that the methylation of the PWS-IC is stable during neuronal lineage conversion, and that the maternal PWS genes remain silenced in PWS NPCs and neurons. Multiple small molecule inhibitors of G9a activate maternal PWS genes in a dose dependent manner in both NPCs and neurons. In addition, G9a inhibitors induce GNRH1 and HTR2C, two neuronal specific genes that contribute to PWS pathology in neurons. Interestingly, distinct from 5-Azacytidine, G9a inhibition does not induce methylation changes of the maternal PWS-IC, indicating that disruption of the histone repressive complex alone is sufficient to drive an open chromatin state at the PWS-IC that leads to partial reactivation of PWS genes.HighlightsModeling PWS disease in a dish using patient derived NPCs and neuronsG9a inhibition activates maternal PWS genes in patient-derived neural cellsG9a inhibition activates maternal SNORD116 and other PWS genes in patient-derived neuronsInhibition of G9a induces PWS downstream genes GNRH1 and HTR2C in PWS neurons

Prader-Willi syndrome: reflections on seminal studies and future therapies

“…These studies revealed that the inhibition of G9a leads to decrease in H3K9me2 marks but no changes to DNA methylation at the PWS-IC. A preprint from Wu et al confirmed the ability of G9a to activate maternal SNORD116 in neural progenitors and neurons derived from human PWS iPSCs [171]. These results probably indicate that G9a plays an important role in the establishment of DNA methylation at the PWS-IC but the activation of the maternal genes are dependent on the status of H3K9 methylation, independent of the PWS-IC DNA methylation.…”

Epigenetics in Prader-Willi Syndrome

“…Specifically, the histone H3 lysine 9 (H3K9) methyltransferase SETDB1 associates with the transcription factor ZNF274 bound to sites within the 5' cluster of SNORD116 repeats, resulting in the deposition of maternal-specific H3K9me3 marks ( Cruvinel et al, 2014 ). Knockdown or inhibition of either SETDB1 or ZNF274 was sufficient to induce a low level of SNORD116 transcript expression from the normally silent maternal allele ( Cruvinel et al, 2014 ; Wu et al, 2019 ; Langouët et al, 2020 ). Together, these results suggest some promise for possible epigenetic therapies that will be discussed at the end of this review.…”