Regulation of jasmonate biosynthesis by posttranslational modification of oxophytodienoate reductase (OPR) 3
Organismic Interactions, Chemical Ecology and Microbiomes of Plant Systems
Final Report Abstract
The induction of plant defense responses against insect herbivores is mediated by jasmonates (JAs) which start to accumulate within minutes after insect attack or wounding. How the rapid formation of these phytohormones is initiated, remains an important open question in the defense signaling field. Rapid accumulation of JAs cannot be explained by changes in gene expression, but calls for post-translational mechanisms of regulation. According to our working hypothesis, post-translational regulation operates at the level of 12-oxophytodienoate reductase (OPR3), a central enzyme of JA biosynthesis. A possible mechanism of regulation was suggested by the crystal structure of OPR3. Tomato OPR3 was found to crystallize as a self-inhibited homo-dimer. The two protomers of the dimer are hooked together by a finger-like structure protruding from each of the subunits, and reaching into the active site of the other. The dimer is stabilized by numerous contacts between the finger of one, and the active site residues of the second protomer. In addition, a sulfate ion located at the dimer interface forms hydrogen bonds with both subunits thus stabilizing the interaction. Intriguingly, the sulfate is located close to the hydroxyl group of Tyr364 (Tyr365 in Arabidopsis) and is positioned perfectly to mimic a phosphorylated tyrosine residue. As a working hypothesis we therefore proposed that the activity of OPR3 is regulated by reversible tyrosine phosphorylation: In healthy plants OPR3 is phosphorylated thus forming the inactive dimer. In response to wounding OPR3 is dephosphorylated, the dimer falls apart, and JA biosynthesis ensues. The goal of this project was to substantiate this hypothesis by showing that OPR3 dimerizes in vivo and that dimerization and inactivation of OPR3 is regulated by phosphorylation of Tyr365 in vivo. Strong evidence was obtained in support of the first part of our hypothesis: Using multiple approaches we could show that OPR3 dimerizes in vivo and that the potentially phosphorylated Tyr365 is required for dimerization. We could also show that the oligomeric state of OPR3 is linked to JA demand: During early stages of flower development, when JAs are needed for anther filament elongation and pollen development, OPR3 is monomeric. When JAs are no longer needed during later stages of flower development, OPR is detected as the inactive dimer. These results are consistent with the proposed regulation of OPR3 activity by reversible dimerization, at least in the developmental context of anther development. However, support is still lacking for the second and essential part of our working hypothesis: Despite considerable efforts using multiple MS/MS approaches, we so far were unable to demonstrate phosphorylation of Tyr365 in vivo.
Publications
- (2012) Regulation der Jasmonsäure-Synthese durch reversible Dimerisierung der Oxophytodiensäurereduktase 3 (OPR3). Dissertation, Universität Hohenheim; Cuvillier Verlag, Göttingen; ISBN 978-3-95404-162-6
Nadja Sperling
- (2013) Regulation der Jasmonsäurebiosynthese durch posttranslationale Modifikation der Oxophytodiensäurereduktase 3 (OPR3). Dissertation, Universität Hohenheim; Cuvillier Verlag, Göttingen; ISBN 978-3-95404-407-8
Kristin Mayer
- (2015) Targeted Analysis of Protein Phosphorylation by 2D Electrophoresis. In Plant Phosphoproteomics: Methods and Protocols (Ed. W. Schulze) Methods Mol. Biol. Vol. 1306: 167-176, Springer Science+Business media, New York
Mayer, K., Albrecht, S., Schaller, A.
(See online at https://doi.org/10.1007/978-1-4939-2648-0_13) - (2017) Regulation of jasmonate biosynthesis and relevance of jasmonates and other phytohormones for resistance against Tomato Yellow Leaf Curl Virus (TYLCV). Dissertation, Universität Hohenheim; Cuvillier Verlag, Göttingen; ISBN 978-3-7369-9694-6
Stephanie Luithardt
- (2018): An ORP3- independent pathway uses 4,5-didehydro-jasmonate for jasmonate synthesis. Nature Chem. Biol. 14: 171–178
Chini, A., Monte, I., Zamarreño, A.M., Hamberg, M., Lassueur, S., Reymond, P., Weiss, S., Stintzi, A., Schaller, A., Porzel, A., García-Mina, J.M., Solano, R.
(See online at https://doi.org/10.1038/NCHEMBIO.2540)
