Breaking the <scp>DNA</scp>‐binding code of <i>Ralstonia solanacearum</i> TAL effectors provides new possibilities to generate plant resistance genes against bacterial wilt disease

Lange, Orlando de; Schreiber, Tom; Schandry, Niklas; Radeck, Jara; Braun, Karl Heinz; Koszinowski, Julia; Heuer, Holger; Strauß, Annett; Lahaye, Thomas

doi:10.1111/nph.12324

Cited by 93 publications

(135 citation statements)

References 46 publications

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“…dTALE UPT AvrBs3 3x Bat1 rep2/6/8 /17 were created as previously described (10) with trimers synthesized by GenScript with the sequences listed in Supplementary Figure S14, while dTALE UPT AvrBs3 3x NI/NN/NG were created with the aforementioned TALE assembly toolkit (15). Repeat domains were assembled into pENTR-D-TALE Δrep Bpi I-AC (15) and then dTALEs transferred into T-DNA binary vector pGWB641 (16) via LR recombination (Life Technologies).…”

Section: Methodsmentioning

confidence: 99%

“…Work on the TALE-like proteins of Ralstonia solanacearum , termed RipTALs, has revealed that they too act as eukaryotic transcription factors and that RipTAL target specificity is linked to RVDs as in TALEs (10). Comparative analysis of TALE and RipTAL repeat arrays also revealed functional differences, due to non-RVD polymorphisms, which could be used to improve custom TALE repeat arrays.…”

Section: Introductionmentioning

confidence: 99%

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Programmable DNA-binding proteins from Burkholderia provide a fresh perspective on the TALE-like repeat domain

Lange

Wolf

Dietze

et al. 2014

Nucleic Acids Research

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The tandem repeats of transcription activator like effectors (TALEs) mediate sequence-specific DNA binding using a simple code. Naturally, TALEs are injected by Xanthomonas bacteria into plant cells to manipulate the host transcriptome. In the laboratory TALE DNA binding domains are reprogrammed and used to target a fused functional domain to a genomic locus of choice. Research into the natural diversity of TALE-like proteins may provide resources for the further improvement of current TALE technology. Here we describe TALE-like proteins from the endosymbiotic bacterium Burkholderia rhizoxinica, termed Bat proteins. Bat repeat domains mediate sequence-specific DNA binding with the same code as TALEs, despite less than 40% sequence identity. We show that Bat proteins can be adapted for use as transcription factors and nucleases and that sequence preferences can be reprogrammed. Unlike TALEs, the core repeats of each Bat protein are highly polymorphic. This feature allowed us to explore alternative strategies for the design of custom Bat repeat arrays, providing novel insights into the functional relevance of non-RVD residues. The Bat proteins offer fertile grounds for research into the creation of improved programmable DNA-binding proteins and comparative insights into TALE-like evolution.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Programmable DNA-binding proteins from Burkholderia provide a fresh perspective on the TALE-like repeat domain

Lange

Wolf

Dietze

et al. 2014

Nucleic Acids Research

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“…The modular DNA recognition domain of transcription activator-like effectors (TALEs) was originally found in natural transcription factors encoded by pathogenic bacteria of the genus Xanthomonas (10,11) and more recently in Ralstonia solanacearum (12). Xanthomonas TALEs are the most widely used in the genome engineering field.…”

Section: Introductionmentioning

confidence: 99%

TALENs facilitate targeted genome editing in human cells with high specificity and low cytotoxicity

Mussolino

Alzubi

Fine

et al. 2014

Nucleic Acids Research

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167

157

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Designer nucleases have been successfully employed to modify the genomes of various model organisms and human cell types. While the specificity of zinc-finger nucleases (ZFNs) and RNA-guided endonucleases has been assessed to some extent, little data are available for transcription activator-like effector-based nucleases (TALENs). Here, we have engineered TALEN pairs targeting three human loci (CCR5, AAVS1 and IL2RG) and performed a detailed analysis of their activity, toxicity and specificity. The TALENs showed comparable activity to benchmark ZFNs, with allelic gene disruption frequencies of 15–30% in human cells. Notably, TALEN expression was overall marked by a low cytotoxicity and the absence of cell cycle aberrations. Bioinformatics-based analysis of designer nuclease specificity confirmed partly substantial off-target activity of ZFNs targeting CCR5 and AAVS1 at six known and five novel sites, respectively. In contrast, only marginal off-target cleavage activity was detected at four out of 49 predicted off-target sites for CCR5- and AAVS1-specific TALENs. The rational design of a CCR5-specific TALEN pair decreased off-target activity at the closely related CCR2 locus considerably, consistent with fewer genomic rearrangements between the two loci. In conclusion, our results link nuclease-associated toxicity to off-target cleavage activity and corroborate TALENs as a highly specific platform for future clinical translation.

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“…The C-terminal region carries conserved three monopartite nuclear localization signals and a conserved eukaryotic-like acidic transcriptional activation domain [6, 17–19]. Notably, TALE-like proteins were also identified in the plant pathogenic bacterium Ralstonia solanacearum [20, 21] offering additional options for engineering DBDs.…”

Section: Tales—virulence Factors Of Xanthomonasmentioning

confidence: 99%

TAL effectors: tools for DNA Targeting

Jankele¹,

Svoboda²

2014

Briefings in Functional Genomics

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Xanthomonas phytopathogenic bacteria produce unique transcription activator-like effector (TALE) proteins that recognize and activate specific plant promoters through a set of tandem repeats. A unique TALE-DNA-binding code uses two polymorphic amino acids in each repeat to mediate recognition of specific nucleotides. The order of repeats determines effector’s specificity toward the cognate nucleotide sequence of the sense DNA strand. Artificially designed TALE-DNA-binding domains fused to nuclease or activation and repressor domains provide an outstanding toolbox for targeted gene editing and gene regulation in research, biotechnology and gene therapy. Gene editing with custom-designed TALE nucleases (TALENs) extends the repertoire of targeted genome modifications across a broad spectrum of organisms ranging from plants and insect to mammals.

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Breaking the DNA‐binding code of Ralstonia solanacearum TAL effectors provides new possibilities to generate plant resistance genes against bacterial wilt disease

Cited by 93 publications

References 46 publications

Programmable DNA-binding proteins from Burkholderia provide a fresh perspective on the TALE-like repeat domain

Programmable DNA-binding proteins from Burkholderia provide a fresh perspective on the TALE-like repeat domain

TALENs facilitate targeted genome editing in human cells with high specificity and low cytotoxicity

TAL effectors: tools for DNA Targeting

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