Abstract
Members of the ancient family of Argonaute (Ago) proteins are present in all domains of life. The common feature of Ago proteins is the ability to bind small nucleic acid guides and use them for sequence-specific recognition-and sometimes cleavage-of complementary targets. While eukaryotic Ago (eAgo) proteins are key players in RNA interference and related pathways, the properties and functions of these proteins in archaeal and bacterial species have just started to emerge. We undertook comprehensive exploration of prokaryotic Ago (pAgo) proteins in sequenced genomes and revealed their striking diversity in comparison with eAgos. Many pAgos contain divergent variants of the conserved domains involved in interactions with nucleic acids, while having extra domains that are absent in eAgos, suggesting that they might have unusual specificities in the nucleic acid recognition and cleavage. Many pAgos are associated with putative nucleases, helicases, and DNA binding proteins in the same gene or operon, suggesting that they are involved in target processing. The great variability of pAgos revealed by our analysis opens new ways for exploration of their functions in host cells and for their use as potential tools in genome editing.IMPORTANCE The eukaryotic Ago proteins and the RNA interference pathways they are involved in are widely used as a powerful tool in research and as potential therapeutics. In contrast, the properties and functions of prokaryotic Ago (pAgo) proteins have remained poorly understood. Understanding the diversity and functions of pAgos holds a huge potential for discovery of new cellular pathways and novel tools for genome manipulations. Only few pAgos have been characterized by structural or biochemical approaches, while previous genomic studies discovered about 300 proteins in archaeal and eubacterial genomes. Since that time the number of bacterial strains with sequenced genomes has greatly expanded, and many previously sequenced genomes have been revised. We undertook comprehensive analysis of pAgo proteins in sequenced genomes and almost tripled the number of known genes of this family. Our research thus forms a foundation for further experimental characterization of pAgo functions that will be important for understanding of the basic biology of these proteins and their adoption as a potential tool for genome engineering in the future.
Highlights
Members of the ancient family of Argonaute (Ago) proteins are present in all domains of life
All eukaryotic Ago (eAgo) and all prokaryotic Ago (pAgo) except one that were experimentally characterized to date possess four domains that are organized in a bilobal structure, with N- and PAZ (PIWI-ArgonauteZwille) domains forming one lobe and MID (Middle) and PIWI (P-element Induced Wimpy Testis) domains forming another lobe [16, 18, 20, 29,30,31,32,33,34]
The genomes of 57 strains carry two pAgo genes, four strains carry three pAgo genes, and one strain encodes four pAgos. pAgos are in general randomly distributed in different prokaryotic clades as the number mbio.asm.org 3 of genera in each class that encode pAgos in their genomes correlates with the total number of genera in the class (Fig. 1A; Fig. S1)
Summary
Members of the ancient family of Argonaute (Ago) proteins are present in all domains of life. Structural and biochemical studies of prokaryotic Agos (pAgos) have provided key insights into the mechanisms of RNAi in eukaryotes and revealed that Ago proteins directly bind short nucleic acid guides and can cleave complementary targets [13,14,15,16,17,18,19,20,21]. The same studies showed that pAgos can associate with short DNA guides and preferentially recognize DNA targets, in contrast to all known eAgos [17, 20, 22,23,24,25,26,27,28] Despite these differences, solved structures of several pAgos and eAgos combined with their sequence alignments revealed a conserved domain organization of these proteins (reviewed in references 12 and 59). Beyond understanding functions of pAgos in their host cells, analysis of pAgos may yield new proteins that can be potentially harnessed for biotechnology, in particular as an alternative to the CRISPR/Cas genome editing tools [42]
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