Abstract
Dicer is a key component of RNA interference (RNAi) and well-known for its role in biogenesis of micro (mi)RNA in the cytoplasm. Increasing evidence suggests that mammalian Dicer is also present and active in the nucleus. We have previously shown that phosphorylated human Dicer associates with chromatin in response to DNA damage and processes double-stranded (ds)RNA in the nucleus. However, a recent study by Much et al. investigated endogenously tagged HA-Dicer both in primary mouse embryonic fibroblasts (PMEFs) as well as adult homozygous viable and fertile HA-Dicer mice under physiological conditions and concluded that murine Dicer is exclusively cytoplasmic. The authors challenged several findings, reporting functions of Dicer in mammalian nuclei. We have re-investigated this issue by applying subcellular fractionation, super-resolution microscopy followed by 3D reconstitution, and phospho-Dicer-specific antibodies using the same HA-Dicer PMEF cell line. Our data show that a small fraction of the murine HA-Dicer pool, approximately 5%, localises in the nucleus and is phosphorylated upon DNA damage. We propose that Dicer localisation is dynamic and not exclusively cytoplasmic, particularly in cells exposed to DNA damage.
Highlights
The endoribonuclease Dicer recognises and processes double-strandedRNA substrates of various origins into small non-codingRNA [1]
Employing the same cells as used by Much and colleagues, we combined super-resolution microscopy followed by 3D reconstitution and biochemical assays to show that endogenously tagged HA-Dicer prominently localises in the nucleus under physiological conditions
We demonstrate that DNA damage triggers accumulation of phosphorylated HA-Dicer in the nucleus, confirming previous observations
Summary
The endoribonuclease Dicer recognises and processes double-stranded (ds)RNA substrates of various origins into small non-coding (nc)RNA [1]. Nuclear Dicer fosters termination of RNAPII transcription [11] and alternative polyadenylation at a subset of protein-coding genes [12] The latter two studies conclude that Dicer association with chromatin may be mediated by the localised production of dsRNA, which is processed into endogenous small interfering (endo-si)RNA to mediate heterochromatin formation by recruitment of G9a methyltransferase in a Dicer-dependent manner. Two studies point toward Dicer-dependent nuclear RNAi in mammals by demonstrating that nuclear, chromatin-associated Dicer impairs expression of the microtubule-binding protein Doublecortin in mouse adult neural stem cells [14] and transactivation of the human secreted frizzled-related protein 1 promoter in cholangiocarcinoma cells [15] These data indicate that Dicer may be present and active in mammalian nuclei to regulate expression of protein-coding genes by both miRNA-dependent and -independent mechanisms
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