RNAi Knockdown of in Maternal Expression of Prader-Willi Syndrome Genes.

BACKGROUND/OBJECTIVES: Euchromatic histone lysine methyltransferase 2 (EHMT2, also known as G9a) is a mammalian histone methyltransferase that catalyzes the dimethylation of histone 3 lysine 9 (H3K9). On human chromosome 15, the parental-specific expression of Prader-Willi Syndrome (PWS)-related genes, such as and , are regulated through the genetic imprinting of the PWS imprinting center (PWS-IC). On the paternal allele, PWS genes are expressed whereas the epigenetic maternal silencing of PWS genes is controlled by the EHMT2-mediated methylation of H3K9 in PWS-IC. Here, we measured the effects of RNA interference of EHMT2 on the maternal expression of genes deficient in PWS in mouse model and patient iPSC-derived cells.

METHODS

We used small interfering RNA (siRNA) oligonucleotides and lentiviral short harpin RNA (shRNA) to reduce / expression in mouse deletion primary neurons, PWS patient-derived induced pluripotent stem cell (iPSC) line and PWS iPSC-derived neurons. We then measured the expression of transcript or protein (if relevant) of PWS genes normally silenced on the maternal allele.

RESULTS

With an approximate reduction of 90% in mRNA and more than 80% of the EHMT2 protein, we demonstrated close to a 2-fold increase in the expression of maternal transcripts for and in PWS iPSCs treated with si compared to PWS iPSC siControl. A similar increase in and RNA expression was observed in PWS iPSC-derived neurons treated with sh.

CONCLUSIONS

RNAi reduction in EHMT2 activates maternally silenced PWS genes. Further studies are needed to determine whether the increase is therapeutically relevant. This study confirms the role of EHMT2 in the epigenetic regulation of PWS genes.