Abstract
Argonaute (Ago) proteins mediate silencing of nucleic acid targets by small RNAs. In fission yeast, Ago1, Tas3 and Chp1 assemble into a RITS complex, which silences transcription near centromeres. Here we describe a repetitive motif within Tas3, termed the 'Argonaute hook', that is conserved from yeast to humans and binds Ago proteins through their PIWI domains in vitro and in vivo. Site-directed mutation of key residues in the motif disrupts Ago binding and heterochromatic silencing in vivo. Unexpectedly, a PIWI domain pocket that binds the 5' end of the short interfering RNA guide strand is required for direct binding of the Ago hook. Moreover, wild-type but not mutant Ago hook peptides derepress microRNA-mediated translational silencing of a target messenger RNA. Proteins containing the conserved Ago hook may thus be important regulatory components of effector complexes in RNA interference.
Publication types
- Research Support, Non-U.S. Gov't
MeSH terms
- Amino Acid Sequence*
- Argonaute Proteins
- Carrier Proteins* / genetics
- Carrier Proteins* / metabolism
- Cell Cycle Proteins / genetics
- Cell Cycle Proteins / metabolism
- Cell Nucleus / metabolism
- Eukaryotic Initiation Factor-2 / genetics
- Eukaryotic Initiation Factor-2 / metabolism
- Gene Silencing
- Humans
- Molecular Sequence Data
- Mutagenesis, Site-Directed
- Protein Binding
- Protein Biosynthesis
- Protein Conformation*
- RNA Interference
- RNA, Small Interfering / genetics
- RNA, Small Interfering / metabolism
- RNA-Binding Proteins
- Schizosaccharomyces / genetics
- Schizosaccharomyces / metabolism
- Schizosaccharomyces pombe Proteins* / chemistry
- Schizosaccharomyces pombe Proteins* / genetics
- Schizosaccharomyces pombe Proteins* / metabolism
- Sequence Alignment
- Two-Hybrid System Techniques
Substances
- AGO2 protein, human
- Ago1 protein, S pombe
- Argonaute Proteins
- Carrier Proteins
- Cell Cycle Proteins
- Chp1 protein, S pombe
- Eukaryotic Initiation Factor-2
- RNA, Small Interfering
- RNA-Binding Proteins
- Schizosaccharomyces pombe Proteins
- Tas3 protein, S pombe