Human Dicer1 hotspot mutation induces both loss and gain of miRNA function

Area of Science:

• Molecular Biology
• Genetics
• Cancer Research

Background:

• Dicer1 is crucial for microRNA (miRNA) biogenesis, with mutations in its RNase IIIb domain frequently observed in cancer.
• Understanding the functional impact of these Dicer1 mutations is limited by the absence of suitable knockin models.

Purpose of the Study:

• To investigate the molecular consequences of Dicer1 mutations, specifically the non-catalytic S1344L variant, on miRNA biogenesis and function.
• To elucidate the role of Dicer1 mutations in altering miRNA strand selection and gene silencing capacity.

Main Methods:

• Generation of Dicer1-knockout (KO) and Dicer1-S1344L mutant human embryonic stem cells (ESCs).
• Utilized in vitro assays, RNA sequencing (RNA-seq), and Argonaute-mRNA profiling to analyze miRNA processing and function.
• Assessed pre-miRNA hairpin processing, strand loading into Argonaute, and subsequent gene repression capacity.

Main Results:

• Dicer1-KO ESCs lacked canonical miRNAs, while S1344L mutants showed ablation of miRNA-5p strands and selective changes in miRNA-3p strands.
• A significant directional upregulation of miRNA-3p passenger strands was observed, indicating a strand-switching phenomenon.
• In vitro assays demonstrated preferential loading of miRNA-3p species from nicked pre-miRNAs into Argonaute, leading to enhanced gene repression.

Conclusions:

• Dicer1 mutations, particularly the S1344L variant, induce a strand switch in miRNA biogenesis, favoring the loading of miRNA-3p strands.
• This altered miRNA processing enhances gene silencing capacity, expanding the known molecular consequences of Dicer1 hotspot mutations in cancer.
• The generated Dicer1 mutant models provide valuable tools for further research into Dicer1-related cancers.