Allelic variation in the indoleacetic acid-lysine synthase gene of the bacterial pathogen Pseudomonas savastanoi and its role in auxin production

Adrián Pintado; Hilario Domínguez-Cerván; Victoria Pastor; Marissa Vincent; Soon Goo Lee; Víctor Flors; Cayo Ramos

doi:10.3389/fpls.2023.1176705

Allelic variation in the indoleacetic acid-lysine synthase gene of the bacterial pathogen Pseudomonas savastanoi and its role in auxin production

Front Plant Sci. 2023 Jun 6:14:1176705. doi: 10.3389/fpls.2023.1176705. eCollection 2023.

Authors

Adrián Pintado^{1

2}, Hilario Domínguez-Cerván^{1

2}, Victoria Pastor³, Marissa Vincent⁴, Soon Goo Lee⁵, Víctor Flors³, Cayo Ramos^{1

2}

Affiliations

¹ Área de Genética, Facultad de Ciencias, Universidad de Málaga (UMA), Málaga, Spain.
² Instituto de Hortofruticultura Subtropical y Mediterránea "La Mayora", Consejo Superior de Investigaciones Científicas (IHSM-UMA-CSIC), Málaga, Spain.
³ Department of Biology, Biochemistry and Natural Sciences, Universitat Jaume I (UJI), Castelló de la Plana, Spain.
⁴ Department of Chemistry and Biochemistry, University of North Carolina Wilmington, Wilmington, NC, United States.
⁵ Department of Molecular and Cellular Biology, Kennesaw State University, Kennesaw, GA, United States.

Abstract

Indole-3-acetic acid (IAA) production is a pathogenicity/virulence factor in the Pseudomonas syringae complex, including Pseudomonas savastanoi. P. savastanoi pathovars (pvs.) genomes contain the iaaL gene, encoding an enzyme that catalyzes the biosynthesis of the less biologically active compound 3-indole-acetyl-ϵ-L-lysine (IAA-Lys). Previous studies have reported the identification of IAA-Lys in culture filtrates of P. savastanoi strains isolated from oleander (pv. nerii), but the conversion of IAA into a conjugate was not detectable in olive strains (pv. savastanoi). In this paper, we show the distribution of iaaL alleles in all available P. savastanoi genomes of strains isolated from woody hosts. Most strains encode two different paralogs, except for those isolated from broom (pv. retacarpa), which contain a single allele. In addition to the three previously reported iaaL alleles (iaaL _Psv, iaaL _Psn and iaaL _Pto), we identified iaaL _Psf, an exclusive allele of strains isolated from ash (pv. fraxini). We also found that the production of IAA-Lys in P. savastanoi pv. savastanoi and pv. nerii depends on a functional iaaL _Psn allele, whereas in pv. fraxini depends on iaaL _Psf. The production of IAA-Lys was detected in cultures of an olive strain heterologously expressing IaaL_Psn-1, IaaL_Psf-1 and IaaL_Psf-3, but not when expressing IaaL_Psv-1. In addition, Arabidopsis seedlings treated with the strains overproducing the conjugate, and thus reducing the free IAA content, alleviated the root elongation inhibitory effect of IAA. IAA-Lys synthase activity assays with purified allozymes confirmed the functionality and specificity of lysine as a substrate of IaaL_Psn-1 and IaaL_Psf-3, with IaaL_Psf-3 showing the highest catalytic efficiency for both substrates. The IAA-Lys synthase activity of IaaL_Psn-1 was abolished by the insertion of two additional tyrosine residues encoded in the inactive allozyme IaaL_Psv-1. These results highlight the relevance of allelic variation in a phytohormone-related gene for the modulation of auxin production in a bacterial phytopathogen.

Keywords: IAA - Indole-3-acetic acid; IAA-lysine synthase; Pseudomonas savastanoi; Pseudomonas syringae; allelic variation; auxin.

Grants and funding

AP and HD-C were supported by the FPU14/05551 and PRE2021-099113 predoctoral grants, respectively. CR was supported by the AGL2017-82492-C2-1-R and PID2020-115177RB-C21 project grants from the Spanish Ministry of Science, Innovation and Universities (Spain), co-financed by the European Regional Development Fund (ERDF). VP and VF were supported by the GV/2018/115 and CIACO/2021/092 grant projects from Generalitat Valenciana and the RTI2018-094350-B-C33 grant from the Spanish Ministry of Science, Innovation and Universities. MV and SL were supported by project grant A20-0079 from The North Carolina Biotechnology Center (NCBC).