CRISPR-Induced Expression of N-Terminally Truncated Dicer in Mouse Cells

RNA interference (RNAi) designates sequence-specific mRNA degradation mediated by small RNAs generated from long double-stranded RNA (dsRNA) by RNase III Dicer. RNAi appears inactive in mammalian cells except for mouse oocytes, where high RNAi activity exists because of an N-terminally truncated Dicer isoform, denoted Dicer^O. Dicer^O processes dsRNA into small RNAs more efficiently than the full-length Dicer expressed in somatic cells. Dicer^O is expressed from an oocyte-specific promoter of retrotransposon origin, which is silenced in other cell types. In this work, we evaluated CRISPR-based strategies for epigenetic targeting of the endogenous Dicer gene to restore Dicer^O expression and, consequently, RNAi. We show that reactivation of Dicer^O expression can be achieved in mouse embryonic stem cells, but it is not sufficient to establish a robust canonical RNAi response.