Evolutionarily conserved roles of the dicer helicase domain in regulating RNA interference processing

Area of Science:

• Molecular Biology
• Enzymology
• Fungal Genetics

Background:

• Dicer enzymes generate small RNAs for gene silencing via RNA interference.
• Dicer helicase domains exhibit varied activities across different Dicer enzymes.
• Evolutionary origins of Dicer helicase functions remain incompletely understood.

Purpose of the Study:

• To investigate the evolutionary origins and functions of Dicer helicase domains.
• To characterize two Dicer enzymes from the thermophilic fungus Sporotrichum thermophile.

Main Methods:

• RNA cleavage assays were performed on S. thermophile Dicer-1 (StDicer-1) and S. thermophile Dicer-2 (StDicer-2).
• ATP hydrolytic activity assays were conducted.
• Functional complementation assays were performed in yeast (Schizosaccharomyces pombe).

Main Results:

• StDicer-1 processed hairpin precursor microRNAs, while StDicer-2 cleaved linear double-stranded RNAs.
• Only StDicer-2 exhibited significant ATP hydrolytic activity with double-stranded RNA.
• Deletion of the StDicer-2 helicase domain enhanced cleavage activity and hairpin RNA affinity.
• Both full-length StDicer-1 and StDicer-2 complemented Dicer-deficient yeast, highlighting helicase domain importance.

Conclusions:

• Fungal Dicer helicase domains possess distinct substrate specificities and regulatory roles.
• The helicase domain is essential for Dicer enzyme function and likely conserved across species.
• These findings suggest an evolutionarily conserved in vivo regulatory role for the Dicer helicase domain.