Repeat 1 of TAL effectors affects target specificity for the base at position zero

Area of Science:

• Plant-microbe interactions
• Molecular biology
• Genetics

Background:

• AvrBs3 is a Xanthomonas transcription-activator-like effector (TALE) that functions as a transcription factor in plant cells.
• TALEs possess DNA-binding domains with repeats that specify DNA base recognition via repeat variable diresidues (RVDs).
• Natural TALE target DNA sequences are typically preceded by a thymine (T0), recognized by a tryptophan residue (W232).

Purpose of the Study:

• To investigate the necessity of the T0 DNA base and the conserved tryptophan residue for AvrBs3-mediated gene activation.
• To explore TALE-DNA interactions with varying upstream bases and TALE domain swaps.

Main Methods:

• Testing TALE mutant derivatives on target sequences with all four possible preceding bases.
• Performing domain swaps between AvrBs3 and TalC from a rice pathogenic Xanthomonas.
• Analyzing the impact of T0 and W232 on gene activation and specificity.

Main Results:

• The T0 DNA base is optimal for TALE function, with its specificity influenced by repeat number and RVD composition.
• The tryptophan residue (W232) and T0 are crucial, particularly when the first repeat's RVD is HD.
• Domain swaps provided insights into the role of specific residues and upstream sequences in TALE targeting.

Conclusions:

• TALE proteins exhibit specific recognition mechanisms for the T0 DNA base, involving repeat number, RVD composition, and conserved residues like W232.
• Understanding these interactions is vital for advancing TALE-based biotechnological applications.
• The findings elucidate novel aspects of TALE-DNA binding and gene regulation in plant-pathogen systems.