Ready to start? Insights on the initiation of the jasmonic acid burst

The biosynthesis of jasmonates, a group of regulators of plant defenses that includes jasmonic acid (JA) and its precursors and derivatives, begins when lipids are freed from the cell membranes of damaged leaves by phospholipase A-Type 1 (PLA1) lipases, such as DEFECTIVE IN ANTHER DEHISCENCE 1 (DAD1) (Wasternack and Strnad, 2018). The product of this reaction is alpha-linoleic acid (α-LA, 18:3) that is further metabolized to 12-oxophytodienoic acid (OPDA). The first steps take place in the plastids and then OPDA is transported to the peroxisomes where it is converted into JA (reviewed by Wasternack and Strnad, 2018).

The biosynthesis of JA from α-LA is a fast process: in response to wounding, the levels of JA dramatically rise within minutes, a phenomenon known as the “JA burst” (Glauser et al., 2008; Koo et al., 2009). Upon the perception of jasmonates, the JA signaling machinery turns on the expression of a wide set of genes, including genes that encode enzymes related to JA biosynthesis: this observation has led to the proposal that JA biosynthesis is regulated by a positive feedback mechanism (Pauwels et al., 2009) and that the JA burst could require the de novo production of JA biosynthetic enzymes. However, multiple reports argue against this hypothesis and definitive evidence supporting positive feedback regulation of JA biosynthesis is lacking (Koch et al., 1999; Miersch and Wasternack, 2000; Pluskota et al., 2007; Koo et al., 2009; Scholz et al., 2015).

In this issue of Plant Physiology, Kimberlin et al. (2022) further refute the hypothesis of JA-upregulated JA biosynthesis by means of a remarkable set of experiments, including approaches from genetics, physiology, pharmacology, biochemistry, and molecular biology. Their results show that wounding, but not JA, enhances the stability of DAD1, and that the availability of its product, α-LA, regulates the onset of JA biosynthesis.

To explore the possibility of positive feedback-enhanced JA biosynthesis, the authors analyzed the accumulation of JA in response to wounding in Arabidopsis (Arabidopsis thaliana) plants pre-treated with inhibitors of transcription or translation in comparison with control plants treated with a mock solution. The JA burst did not change between treatments, as JA accumulated to a similar level in all cases. Then, the authors isolated intact chloroplasts from pea (Pisum sativum) leaves and fed them with α-LA, leading to a dramatic production of OPDA that saturated within 5 min, supporting the idea that α-LA can activate JA synthesis (Vick and Zimmerman, 1983; Farmer and Ryan, 1992; Christeller and Galis, 2014). As all JA biosynthetic enzymes are encoded by nuclear genes, the feeding experiment suggests that the biosynthesis machinery was already present and functional in the chloroplasts. Together, these results consistently confirm that JA burst is independent of the transcription or translation of de novo enzymes and that α-LA is the limiting factor at the initiation of JA production.

If the entire set of biosynthetic genes were under the control of a positive feedback mechanism, any minor stress or developmental program that raises JA levels could cause a never-ending loop in JA production. To set aside this auto-amplification theory, the authors manipulated the expression of DAD1, a gene that encodes an enzyme that catalyzes the production of α-LA. The transcription of DAD1 is upregulated by mechanical wounding, although, in contrast to other JA biosynthetic genes, it cannot be enhanced by JA treatment. When DAD1 was expressed under the promoter of OXOPHYTODIENOATE-REDUCTASE 3 (OPR3), whose expression is upregulated by JA, plants accumulated high levels of JA even in resting conditions. Furthermore, the OPR3pro:DAD1 lines were severely stunted and accumulated anthocyanins (Figure 1), a phenotype reminiscent of mutants with constitutive JA signaling (Campos et al., 2016; Guo et al., 2018). These observations illustrate the consequences of having DAD1 expression controlled by JA and the detrimental impact derived from perpetual JA biosynthesis.

Figure 1.

Transgenic Arabidopsis expressing DAD1 controlled by JA-responsive promoter displays symptoms of chronic JA accumulation. Photographs of 4-week-old wild-type (WT), OPR3pro:DAD1 (three independent lines) and OPR3pro:DAD1 in allene oxide synthase (aos) background grown under normal growth condition. AOS is a JA biosynthetic gene and aos knockout lines cannot produce JA (Park et al., 2002). Adapted from Kimberlin et al. (2022).

Wounding-enhanced DAD1 expression lags beyond the quick JA burst, and as explained previously, the JA burst does not require de novo synthesis of enzymes. So, the question arises: how does wounding regulate DAD1 at the beginning of JA biosynthesis? To answer this question, the authors engineered a DAD1-Myc construction expressed under a dexamethasone (dex)-inducible promoter. After dex treatment, DAD1 protein was detected using anti-Myc antibodies. If an inhibitor of translation was applied after dex induction, DAD1 was quickly turned over, suggesting this enzyme is highly unstable in resting conditions. However, when plants were wounded, DAD1 was protected from degradation and remained at a high level for a longer time after the inhibitor treatment compared with unwounded controls. In agreement, when the dex:DAD1-Myc lines were treated with a combination of dex and wounding, JA accumulated more than with the separate treatments.

In summary, the results of Kimberlin et al. (2022) present further evidence against positive feedback-regulated JA biosynthesis. Although JA enhances the expression of several JA biosynthetic genes, the de novo production of enzymes is not required at the onset of JA synthesis. DAD1, an enzyme that produces α-LA and acts as the limiting factor in the beginning of JA synthesis, is highly unstable under resting conditions but could be stabilized by wounding, suggesting a possible post-translational mechanism by which mechanical wounding but not JA can control the onset of JA biosynthesis. This mechanism may prevent never-ending positive feedback in JA production, which could cause severe growth abnormalities, as evidenced by the phenotype of the promOPR3:DAD1 transgenic plants. Future research may focus on how wounding prevents DAD1 degradation. Here, Kimberlin and collaborators have set the starting point.

Conflict of interest statement. None declared.