Ectopic Expression ofAtJMTinNicotiana attenuata: Creating a Metabolic Sink Has Tissue-Specific Consequences for the Jasmonate Metabolic Network and Silences Downstream Gene Expression    

Abstract

To create a metabolic sink in the jasmonic acid (JA) pathway, we generated transgenic Nicotiana attenuata lines ectopically expressing Arabidopsis (Arabidopsis thaliana) jasmonic acid O-methyltransferase (35S-jmt) and additionally silenced in other lines the N. attenuata methyl jasmonate esterase (35S-jmt/ir-mje) to reduce the deesterification of methyl jasmonate (MeJA). Basal jasmonate levels did not differ between transgenic and wild-type plants; however, after wounding and elicitation with Manduca sexta oral secretions, the bursts of JA, jasmonoyl-isoleucine (JA-Ile), and their metabolites that are normally observed in the lamina, midvein, and petiole of elicited wild-type leaves were largely absent in both transformants but replaced by a burst of endogenous MeJA that accounted for almost half of the total elicited jasmonate pools. In these plants, MeJA became a metabolic sink that affected the jasmonate metabolic network and its spread to systemic leaves, with major effects on 12-oxo-phytodieonic acid, JA, and hydroxy-JA in petioles and on JA-Ile in laminas. Alterations in the size of jasmonate pools were most obvious in systemic tissues, especially petioles. Expression of threonine deaminase and trypsin proteinase inhibitor, two JA-inducible defense genes, was strongly decreased in both transgenic lines without influencing the expression of JA biosynthesis genes that were uncoupled from the wounding and elicitation with M. sexta oral secretions-elicited JA-Ile gradient in elicited leaves. Taken together, this study provides support for a central role of the vasculature in the propagation of jasmonates and new insights into the versatile spatiotemporal characteristics of the jasmonate metabolic network.

Silencing Nicotiana attenuata Calcium-Dependent Protein Kinases, CDPK4 and CDPK5, Strongly Up-Regulates Wound- and Herbivory-Induced Jasmonic Acid Accumulations

“…To examine this possibility, IRcdpk4/5-1 was crossed with IRcoi1 (silenced in COI1, the receptor for JA-Ile; Paschold et al, 2007) to abolish JA signaling (IRcdpk4/ 5-1 3 IRcoi1 plants). Moreover, IRcdpk4/5-1 line was also crossed with a line ectopically expressing an Arabidopsis JASMONIC ACID CARBOXYL METH-YLTRANSFERASE (ovJMT; Seo et al, 2001), which converts JA to methyl jasmonate, an inactive form of jasmonate in N. attenuata (Stitz et al, 2011;IRcdpk4/ 5- 1 3 ovJMT plants). Notably, the morphologies of both IRcdpk4/5-1 3 IRcoi1 and IRcdpk4/5-1 3 ovJMT plants were indistinguishable from those of IRcoi1 and ovJMT, respectively, including having similar stem lengths to those of IRcoi1 and ovJMT, normal leaf shape, and partial male sterility due to deficiencies in JA signaling and JA accumulation (M. Heinrich, C. Hettenhausen, I.T.…”

Navigating natural variation in herbivory-induced secondary metabolism in coyote tobacco populations using MS/MS structural analysis

“…One treated leaf per plant was harvested 1 h after treatment and analyzed for jasmonates as described in ref. 61.…”

Empoasca leafhoppers attack wild tobacco plants in a jasmonate-dependent manner and identify jasmonate mutants in natural populations

“…The analysis of OPDA, JA, JA-Ile, MeJA, JA-Val, 11/12-hydroxy-JA, 11/12-hydroxy-JA-Ile, and 11/12-carboxy-JA-Ile was performed as previously described (29,47). The PCA (Fig.…”

“…1A). Additionally, we ectopically expressed JA methyl transferase 1 (JMT1; 35S-jmt1) to deplete jasmonate accumulation metabolically by redirecting the flux of JA into methyl-JA (MeJA) (47). Compared with control plants, 35S-jmt1 plants have reduced levels of JA-Ile after simulated herbivory, and therefore their COI1-mediated JA-signaling capacity is reduced (47).…”