TAL nucleases (TALNs): hybrid proteins composed of TAL effectors and FokI DNA-cleavage domain

By Tingyu Li, Sheng Huang, W. Jiang, David A. Wright, M. Spalding, D. Weeks, Bing Yang. Nucleic Acids Res., 2010

548 citations recorded by the source corpus.

DNA double-strand breaks enhance homologous recombination in cells and have been exploited for targeted genome editing through use of engineered endonucleases. Here we report the creation and initial characterization of a group of rare-cutting, site-specific DNA nucleases produced by fusion of the restriction enzyme FokI endonuclease domain (FN) with the high-specificity DNA-binding domains of AvrXa7 and PthXo1. AvrXa7 and PthXo1 are members of the transcription activator-like (TAL) effector family whose central repeat units dictate target DNA recognition and can be modularly constructed to create novel DNA specificity. The hybrid FN-AvrXa7, AvrXa7-FN and PthXo1-FN proteins retain both recognition specificity for their target DNA (a 26 bp sequence for AvrXa7 and 24 bp for PthXo1) and the double-stranded DNA cleaving activity of FokI and, thus, are called TAL nucleases (TALNs). With all three TALNs, DNA is cleaved adjacent to the TAL-binding site under optimal conditions in vitro. When expressed in yeast, the TALNs promote DNA homologous recombination of a LacZ gene containing paired AvrXa7 or asymmetric AvrXa7/PthXo1 target sequences. Our results demonstrate the feasibility of creating a tool box of novel TALNs with potential for targeted genome modification in organisms lacking facile mechanisms for targeted gene knockout and homologous recombination.

Subfields: Molecular Biology, Plant Science

View Paper: https://www.semanticscholar.org/paper/2692b1fcb42809c17fe63bae53c2bf359b71b25c

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Citations in the corpus

Genome Engineering With Zinc-Finger Nucleases (2011, background): “Fusions of TAL domains to the FokI nuclease domain (TALNs or TALENs) direct cleavage to specific sites both in vitro and in vivo (Christian et al. 2010; Li et al. 2011; Miller et al. 2011).”
A CRISPR/Cas9 Toolbox for Multiplexed Plant Genome Editing and Transcriptional Regulation1[OPEN] (2015, background): “971 surfaced, a more versatile transcription activator-like effector nuclease (TALEN)-based SSN platform emerged (Christian et al., 2010; Li et al., 2011; Miller et al., 2011).”
A Transcription Activator-Like Effector (TALE) Toolbox for Genome Engineering (2012, background): “When coupled to effector domains, customized TALEs provide a promising platform for achieving a wide variety of targeted genome manipulation”
A TALE nuclease architecture for efficient genome editing (2011, background): “These results have spurred interest in the potential use of TALE-nuclease chimeras (TALENs”
Delivering CRISPR: a review of the challenges and approaches (2018, background): “Shortly after the discovery of TALEs, TALE nucleases (TALENs) were developed that, like ZFNs, are a fusion protein comprised of a TALE and a FokI nuclease (Christian et al., 2010; Miller et al., 2011; Li et al., 2011; Reyon et al., 2012).”
A novel TALE nuclease scaffold enables high genome editing activity in combination with low toxicity (2011, methodology): “While TALENs with long linkers (>200 amino acids) seem to work on a very broad range of spacers spanning 14–40 bp (29,30), the shorter linkers (17–63 residues) used in our study and in Miller et al.”
TAL Effector-Nucleotide Targeter (TALE-NT) 2.0: tools for TAL effector design and target prediction (2012, methodology): “In the case of TALENs, the optimum spacer length between the TALEN monomers depends on the architecture used (5,13,15,24).”
Gene therapy comes of age (2018, unknown): “The demonstration in 2009 that the DNA binding domain of bacterial proteins called transcription activator–like effectors (TALEs) can be readily altered (47, 48) opened the door to the creation of TALE nucleases (TALENs) (49, 50).”
Genome Engineering of Crops with Designer Nucleases (2012, background): “, 2012), yeast (Saccharomyces cerevisiae) (Li et al., 2011), zebrai sh (Huang et al.”
Efficient design and assembly of custom TALEN and other TAL effector-based constructs for DNA targeting (2011, background): “In particular, we and other groups have shown that TAL effectors can be fused to the catalytic domain of the FokI nuclease to create targeted DNA double-strand breaks (DSBs) in vivo for genome editing (8,10,12,13).”
TALENs: a widely applicable technology for targeted genome editing (2012, unknown): “23–25 More recently, co-crystal structures of TALE DNA-binding domains bound to their cognate sites have shown that individual repeats comprise two-helix v-shaped bundles that stack to form a superhelix around the DNA and the hypervariable residues at positions 12 and 13 are positioned in the DNA…”