title: Characterization and DNA-binding specificities of Ralstonia TAL-like effectors.
abstract: Transcription activator-like effectors (TALEs) from Xanthomonas sp. have been used as customizable DNA-binding modules for genome-engineering applications. Ralstonia solanacearum TALE-like proteins (RTLs) exhibit similar structural features to TALEs, including a central DNA-binding domain composed of 35 amino acid-long repeats. Here, we characterize the RTLs and show that they localize in the plant cell nucleus, mediate DNA binding, and might function as transcriptional activators. RTLs have a unique DNA-binding architecture and are enriched in repeat variable di-residues (RVDs), which determine repeat DNA-binding specificities. We determined the DNA-binding specificities for the RVD sequences ND, HN, NP, and NT. The RVD ND mediates highly specific interactions with C nucleotide, HN interacts specifically with A and G nucleotides, and NP binds to C, A, and G nucleotides. Moreover, we developed a highly efficient repeat assembly approach for engineering RTL effectors. Taken together, our data demonstrate that RTLs are unique DNA-targeting modules that are excellent alternatives to be tailored to bind to user-selected DNA sequences for targeted genomic and epigenomic modifications. These findings will facilitate research concerning RTL molecular biology and RTL roles in the pathogenicity of Ralstonia spp.
authors.affiliation: Center for Desert Agriculture, King Abdullah University of Science and Technology, Thuwal 23955-6900, Kingdom of Saudi Arabia.
authors.forename: Lixin
authors.initials: L
authors.lastname: Li
authors.forename: Ahmed
authors.initials: A
authors.lastname: Atef
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Piatek
authors.forename: Zahir
authors.initials: Z
authors.lastname: Ali
authors.forename: Marek
authors.initials: M
authors.lastname: Piatek
authors.forename: Mustapha
authors.initials: M
authors.lastname: Aouida
authors.forename: Altanbadralt
authors.initials: A
authors.lastname: Sharakuu
authors.forename: Ali
authors.initials: A
authors.lastname: Mahjoub
authors.forename: Guangchao
authors.initials: G
authors.lastname: Wang
authors.forename: Suhail
authors.initials: S
authors.lastname: Khan
authors.forename: Nina V
authors.initials: NV
authors.lastname: Fedoroff
authors.forename: Jian-Kang
authors.initials: JK
authors.lastname: Zhu
authors.forename: Magdy M
authors.initials: MM
authors.lastname: Mahfouz
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D003597:Cytosine Nucleotides / Q000378:metabolism; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D043367:Ralstonia; D020449:Repetitive Sequences, Amino Acid; D013379:Substrate Specificity; D015533:Transcriptional Activation
chemical_list: D001426:Bacterial Proteins; D003597:Cytosine Nucleotides; D004247:DNA
keywords: Ralstonia solanacearum; TAL effectors; TALE activators and repressors; TALE nucleases (TALENs); genome engineering; targeted genome modifications