Arabidopsis thaliana GH3.5 acyl acid amido synthetase mediates metabolic crosstalk in auxin and salicylic acid homeostasis
Significance Plants require precise control over growth regulators during development and in their responses to biotic and abiotic stresses. One strategy for modulating levels of bioactive phytohormones is to conjugate these molecules to amino acids using acyl acid amido synthetases of the Gretchen Hagen 3 (GH3) protein family. Typically, GH3 proteins modify one type of phytohormone. Structural studies, along with in vitro and in planta biochemical analyses, reveal that the GH3.5 protein from the model plant Arabidopsis thaliana conjugates multiple molecules from various phytohormone pathways. This activity mediates crosstalk between auxin developmental and pathogen response pathways. In Arabidopsis thaliana, the acyl acid amido synthetase Gretchen Hagen 3.5 (AtGH3.5) conjugates both indole 3 acetic acid (IAA) and salicylic acid (SA) to modulate auxin and pathogen response pathways. To understand the molecular basis for the activity of AtGH3.5, we determined the X ray crystal structure of the enzyme in complex with IAA and AMP. Biochemical analysis demonstrates that the substrate preference of AtGH3.5 is wider than originally described and includes the natural auxin phenylacetic acid (PAA) and the potential SA precursor benzoic acid (BA) Residues that determine IAA versus BA substrate preference were identified. The dual functionality of AtGH3.5 is unique to this enzyme although multiple IAA conjugating GH3 proteins share nearly identical acyl acid binding sites. In planta analysis of IAA, PAA, SA, and BA and their respective aspartyl conjugates were determined in wild type and overexpressing lines of A. thaliana. This study suggests that AtGH3.5 conjugates auxins (i.e. IAA and PAA) and benzoates (i.e. SA and BA) to mediate crosstalk between different metabolic pathways, broadening the potential roles for GH3 acyl acid amido synthetases in plants.
Proceedings of the National Academy of Sciences