TAL effectors: Function, structure, engineering and applications
ABSTRACT TAL effectors are proteins secreted by bacterial pathogens into plant cells, where they enter the nucleus and activate expression of individual genes. TAL effectors display a modular architecture that includes a central DNA-binding region comprising a tandem array of nearly identical repeats that are almost all 34 residues long. Residue number 13 in each TAL repeat (one of two consecutive polymorphic amino acids that are termed 'repeat variable diresidues', or 'RVDs') specifies the identity of a single base; collectively the sequential repeats and their RVDs dictate the recognition of sequential bases along one of the two DNA strands. The modular architecture of TAL effectors has facilitated their extremely rapid development and application as artificial gene targeting reagents, particularly in the form of site-specific nucleases. Recent crystallographic and biochemical analyses of TAL effectors have established the structural basis of their DNA recognition properties and provide clear directions for future research.
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ABSTRACT: Transcription activator-like (TAL) effectors are encoded by plantpathogenic bacteria and induce expression of plant host genes. TAL effectors bind DNA on the basis of a unique code that specifies binding of amino acid pairs in repeat units to particular DNA bases in a one-to-one correspondence. This code can be used to predict binding sites of natural TAL effectors and to design novel synthetic DNA-binding domains for targeted genome manipulation. Natural mechanisms of resistance in plants against TAL effector-containing pathogens have given insights into new strategies for disease control. Expected final online publication date for the Annual Review of Phytopathology Volume 51 is August 04, 2013. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.Annual Review of Phytopathology 05/2013; DOI:10.1146/annurev-phyto-082712-102255 · 11.00 Impact Factor
- Molecular Plant 05/2013; 6(5). DOI:10.1093/mp/sst075 · 6.61 Impact Factor
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ABSTRACT: Abstract Synthetic targeted endonucleases such as zinc finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) have recently emerged as powerful tools for targeted mutagenesis, especially in organisms that are not amenable to embryonic stem cell manipulation. Both ZFNs and TALENs consist of DNA-binding arrays that are fused to the nonspecific FokI nuclease domain. In an effort to improve targeted endonuclease mutagenesis efficiency, we enhanced their catalytic activity using the Sharkey FokI nuclease domain variant. All constructs tested display increased DNA cleavage activity in vitro. We demonstrate that one out of four ZFN arrays containing the Sharkey FokI variant exhibits a dramatic increase in mutagenesis frequency in vivo in zebrafish. The other three ZFNs exhibit no significant alteration of activity in vivo. Conversely, we demonstrate that TALENs containing the Sharkey FokI variant exhibit absent or severely reduced in vivo mutagenic activity in zebrafish. Notably, Sharkey ZFNs and TALENs do not generate increased toxicity-related defects or mortality. Our results present Sharkey ZFNs as an effective alternative to conventional ZFNs, but advise against the use of Sharkey TALENs.Zebrafish 06/2013; 10(3). DOI:10.1089/zeb.2012.0832 · 1.77 Impact Factor