Prokaryotic Argonaute Protein from Natronobacterium gregoryi Requires RNAs To Activate for DNA Interference In Vivo
Jiani Xing,
Lixia Ma,
Xinzhen Cheng,
Jinrong Ma,
Ruyu Wang,
Kun Xu,
Joe S. Mymryk,
Zhiying Zhang
Affiliations
Jiani Xing
Key Laboratory of Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi, China
Lixia Ma
Key Laboratory of Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi, China
Xinzhen Cheng
Key Laboratory of Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi, China
Jinrong Ma
Key Laboratory of Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi, China
Ruyu Wang
Key Laboratory of Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi, China
Kun Xu
Key Laboratory of Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi, China
Joe S. Mymryk
Department of Microbiology & Immunology, Oncology and Otolaryngology, The University of Western Ontario, London, Ontario, Canada
Zhiying Zhang
Key Laboratory of Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi, China
ABSTRACT The Argonaute proteins are present in all three domains of life, which are archaea, bacteria, and eukarya. Unlike the eukaryotic Argonaute proteins, which use small RNA guides to target mRNAs, some prokaryotic Argonaute proteins (pAgos) use a small DNA guide to interfere with DNA and/or RNA targets. However, the mechanisms of pAgo natural function remain unknown. Here, we investigate the mechanism by which pAgo from Natronobacterium gregoryi (NgAgo) targets plasmid and bacteriophage T7 DNA using a heterologous Escherichia coli-based model system. We show that NgAgo expressed from a plasmid linearizes its expression vector. Cotransformation assays demonstrate that NgAgo requires an RNA in trans that is transcribed from the bacteriophage T7 promoter to activate cleavage of a cotransformed plasmid, reminiscent of the trans-RNA function in CRISPR/Cas9. We propose a mechanism to explain how NgAgo eliminates invading foreign DNA and bacteriophage. By leveraging this discovery, we show that NgAgo can be programmed to target a plasmid or a chromosome locus. IMPORTANCE We revealed the mechanism that explains how the NgAgo eliminates the invading foreign DNA and bacteriophage in bacterial cells at 37°C, and by leveraging this discovery, NgAgo can be programmed to target a plasmid or a chromosome locus.
