Fig. 1. Phylogenetic tree and crystal structure of the Ago/Piwi proteins. (A) Phylogenetic tree of the Ago/Piwi protein family. Alignments were generated using ClustalW (http://www.ebi.ac.uk/Tools/clustalw/). The length of each branch represents an estimate of the genetic distance. The alignment was done using the PAZ domain (when present), the PIWI domain and the C terminus. The sequences are based mainly on published RefSeqs for PAZ/PIWI domain-containing proteins at PubMed (http://www.ncbi.nlm.nih.gov/sites/entrez). The Accession numbers of the sequences can be obtained from the authors. Asterisks indicate Ago/Piwi members with experimentally determined cleavage activity. The interacting classes of small RNAs are indicated next to their corresponding Ago/Piwi family members (evidence stems from biochemical and/or genetic experiments). Ago/Piwi family members are designated according to their affiliation to either the Ago protein family, the Class 3 protein family or the Piwi protein family. C. elegans Alg1, Alg2, T23D8.7, ZK757.3, T22B3.2 comprise the Ago family; PRG-1, PRG-2 comprise the Piwi family; Sago-1, Sago-2, PPW-1, R06C7.1, F55A12.1, PPW-2, F58G1.1, C06A1.4, R04A9.2, Y49F6A.1, T22H9.3, C16C10.3, CSR-1, M03D4.6, ZK1248.7, C14B1.7, C04F12.1 comprise the Class 3 family. Mammalian Ago proteins are also known as eIF2Cs (eukaryotic translation initiation factors). Mammalian Piwil1 is also known as Hiwi; Miwi or Riwi depending on the species (human or mouse or rat), Piwil2 is also known as Hili, and Piwil4 as Hiwi2. Cniwi is the Piwi protein in Podocoryne carnea, and Seawi is the Piwi member in the sea urchins Strongylocentrotus purpuratus and Paracentrotus lividus. (B) Ribbon diagram of the structure of the Aquifex aeolicus Piwi protein, showing its bilobed architecture. The functions of the protein domains are further discussed in the text.