DICER cleavage fidelity is governed by 5'-end binding pockets

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• RNA interference (RNAi) is crucial for gene regulation, mediated by the DICER enzyme.
• DICER processes RNA precursors into small regulatory RNAs using a 5'-end counting rule.
• Previous models suggested a single 5'-end binding pocket in DICER, potentially causing cleavage inaccuracies.

Purpose of the Study:

• To investigate the role of 5'-end nucleotides in DICER cleavage accuracy.
• To elucidate the structural basis of substrate recognition and cleavage by DICER.
• To expand the mechanistic understanding of small RNA biogenesis in metazoans.

Main Methods:

• Massively parallel dicing assays to analyze DICER activity.
• Cryo-electron microscopy to determine DICER-substrate complex structures.
• Computational modeling to assess RNA conformational changes.

Main Results:

• Identified a conserved guanosine-favored (G-favored) binding pocket distinct from the previously known uridine-favored (U-favored) pocket.
• Demonstrated that 5'-G promotes precise cleavage for many RNA substrates.
• Revealed that dual 5'-end pockets, RNA-motif recognition, and domain motions (dsRBD, PAZ) integrate for accurate cleavage site selection.

Conclusions:

• DICER employs dual 5'-end binding pockets to ensure precise RNA precursor processing.
• RNA-motif recognition and domain dynamics contribute to accurate cleavage site selection, expanding the mechanism of microRNA biogenesis.