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J Biol Chem. 2014 Oct 10;289(41):28352-62. doi: 10.1074/jbc.M114.589051. Epub 2014 Aug 18.

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

Author information

1
From the Department of Molecular and Cell Biology.
2
From the Department of Molecular and Cell Biology, Howard Hughes Medical Institute, and Department of Chemistry, University of California at Berkeley and the Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720 doudna@berkeley.edu.

Abstract

The enzyme Dicer generates 21-25 nucleotide RNAs that target specific mRNAs for silencing during RNA interference and related pathways. Although their active sites and RNA binding regions are functionally conserved, the helicase domains have distinct activities in the context of different Dicer enzymes. To examine the evolutionary origins of Dicer helicase functions, we investigated two related Dicer enzymes from the thermophilic fungus Sporotrichum thermophile. RNA cleavage assays showed that S. thermophile Dicer-1 (StDicer-1) can process hairpin precursor microRNAs, whereas StDicer-2 can only cleave linear double-stranded RNAs. Furthermore, only StDicer-2 possesses robust ATP hydrolytic activity in the presence of double-stranded RNA. Deletion of the StDicer-2 helicase domain increases both StDicer-2 cleavage activity and affinity for hairpin RNA. Notably, both StDicer-1 and StDicer-2 could complement the distantly related yeast Schizosaccharomyces pombe lacking its endogenous Dicer gene but only in their full-length forms, underscoring the importance of the helicase domain. These results suggest an in vivo regulatory function for the helicase domain that may be conserved from fungi to humans.

KEYWORDS:

Dicer; Fungi; MicroRNA Biogenesis; RNA Helicase; RNA Interference (RNAi); Ribonuclease

PMID:
25135636
PMCID:
PMC4192488
DOI:
10.1074/jbc.M114.589051
[Indexed for MEDLINE]
Free PMC Article

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