Flexible TALEs for an expanded use in gene activation, virulence and scaffold engineering

verfasst von

Sebastian Becker, Stefanie Mücke, Jan Grau, Jens Boch

Abstract

Transcription activator-like effectors (TALEs) are bacterial proteins with a programmable DNA-binding domain, which turned them into exceptional tools for biotechnology. TALEs contain a central array of consecutive 34 amino acid long repeats to bind DNA in a simple one-repeat-to-one-nucleotide manner. However, a few naturally occurring aberrant repeat variants break this strict binding mechanism, allowing for the recognition of an additional sequence with a -1 nucleotide frameshift. The limits and implications of this extended TALE binding mode are largely unexplored. Here, we analyse the complete diversity of natural and artificially engineered aberrant repeats for their impact on the DNA binding of TALEs. Surprisingly, TALEs with several aberrant repeats can loop out multiple repeats simultaneously without losing DNA-binding capacity. We also characterized members of the only natural TALE class harbouring two aberrant repeats and confirmed that their target is the major virulence factor OsSWEET13 from rice. In an aberrant TALE repeat, the position and nature of the amino acid sequence strongly influence its function. We explored the tolerance of TALE repeats towards alterations further and demonstrate that inserts as large as GFP can be tolerated without disrupting DNA binding. This illustrates the extraordinary DNA-binding capacity of TALEs and opens new uses in biotechnology.

Organisationseinheit(en)

Abteilung Pflanzenbiotechnologie
Institut für Pflanzengenetik

Externe Organisation(en)

Martin-Luther-Universität Halle-Wittenberg

Typ

Artikel

Journal

Nucleic Acids Research

Band

50

Seiten

2387-2400

Anzahl der Seiten

14

ISSN

0301-5610

Publikationsdatum

28.02.2022

Publikationsstatus

Veröffentlicht

Peer-reviewed

Ja

ASJC Scopus Sachgebiete

Genetik

Elektronische Version(en)

https://doi.org/10.1093/nar/gkac098 (Zugang: Offen)