Insect herbivores selectively mute GLV production in plants

Abstract

Through co-evolution insect herbivores have developed a myriad of strategies to manipulate host plant defense responses that include the synthesis of defensive compounds whose composition depends on the insect feeding mode.  Among the plant-produced compounds are jasmonates (JAs), and Green Leafy Volatiles (GLVs), metabolites produced by the two parallel and competing branches of the oxylipin pathway. Here we provide evidence that chewing insects stimulate JA production but suppress the synthesis of GLVs through the transcriptional and post transcriptional reprogramming of critical genes in the corresponding pathway. We further establish that herbivore-derived elicitors known as Herbivore-Associated Molecular Patterns (HAMPs) are responsible for the reprogramming of these pathway genes. Through this strategy chewing herbivores coerce the plant signaling machinery that would otherwise leads to a reduction in the nutritional quality of the immediate and neighboring plants, and additionally shelters the herbivores from their natural enemies that are otherwise guided by the GLV cues to prey-infested plants. 

Plant volatiles are communication signals that mediate intra-, inter-plant, and plant-insect interactions, and therefore they are key in maintaining ecological homeostasis. These signaling compounds are of diverse chemical nature generated from both primary and secondary metabolites, and their distinct signature, as defined by their quantitative and qualitative values, determines their effectiveness in maintaining the ecological balance.¹

Insects make up the most diverse and abundant group of plant consumers and interaction between plants and insect herbivores is the most frequent known interspecies communication and a driving force of evolution.^2,3 Among several classes of plant metabolites produced in response to insect attack are oxylipins—the oxygenated derivatives of fatty acids generated by parallel and competing branches of oxylipin pathway.^4-7 The best studied oxylipin branches however are allene oxide synthase (AOS) and hydroperoxide lyase (HPL) responsible for production of jasmonates [jasmonic acid (JA), methyl jasmonate (MeJA) and their biosynthetic precursor, 12- oxophytodienoic acid (12-OPDA)], and the green leafy volatiles (GLVs), respectively^8,9 (Fig. 1). Both these pathway metabolites are known to defend plants, directly or indirectly, against insects. Specifically, JA is an indispensable signaling component of the defense pathway that promotes resistance to a wide spectrum of insects,^10-13 and GLVs are known to be involved in intra- and interplant defense signaling cascades and in directing tritrophic (plant-herbivore-natural enemy) interactions.^7,9,14-17

Figure 1. Simplified scheme of oxylipin biosynthesis pathway. Abbreviations: LOXs, lipoxygenases; AOS, allene oxide synthase; HPL, hydroperoxide lyase.

Recently we have examined the influence of chewing insect feeding and mechanical wounding on the HPL- and AOS- derived metabolic signatures, by using three Arabidopsis genotypes, namely Col-0 accession, a natural loss-of-function mutation in hpl¹⁸ as the control, engineered Col-0 lines that overexpress a rice HPL under the control of a 35S promoter (OsHPL3 OE)¹⁹ and wild-type Wassilewskija (Ws) ecotype that contain a functional HPL driven by the native promoter.²⁰ These genotypes were damaged either mechanically or by insects of different feeding guilds (piercing aphids, generalist chewing caterpillars “Spodoptera exicua” and specialist chewing caterpillars “Pieris rapae”). Subsequent metabolic analyses established that emission of GLVs is stimulated by wounding incurred mechanically or by aphids, but the release of these volatiles is constitutively impaired by both generalist and specialist chewing insects (Fig. 2). Simultaneously however, these chewing herbivores stimulated JA production, demonstrating targeted suppression of the HPL branch of the oxylipin pathway. Employment of lines engineered to express HPL constitutively, in conjunction with qRT-PCR-based expression analyses, established a combination of transcriptional and posttranscriptional reprogramming of the HPL-pathway genes as the mechanistic basis of insect-mediated suppression of the corresponding metabolites. Additional studies established that insect-inflicted suppression of GLV emission is caused by herbivore oral secretion (OS), as a strategy to counteract the plant’s ability to produce defensive metabolites and to reduce nutritional quality central to herbivore fitness. Subsequent no-choice feeding trials on plants that had or had not been exposed to GLVs from a neighboring damaged plant determined that caterpillars fed more on plants that had not been primed by GLVs emitted by wounded plants, suggesting that plant-plant communication triggered by these volatiles altered the insect feeding behavior. This insect-mediated suppression of GLV production is also instrumental for the removal of this broad ecological signal that could otherwise attract and direct herbivore’s natural enemies.

Figure 2. Emission of GLVs is stimulated by mechanical wounding, but is suppressed by chewing insects. Gas chromatography-mass spectrometry (GC-MS) ion chromatograms (m/z 67) showing cis-3-hexenol and cis-3-hexenyl acetate in volatile samples collected from control (black line), wounded (red line), and caterpillar damaged (blue line) Arabidopsis plants. Volatiles were captured for 3 h on mini-columns filled with Porapak Q^® 100/120 (Alltech Associates, Inc.) using an open-flow system, bound compounds were eluted with 300 µl of dichloromethane and analyzed by GC-MS.

These findings reveal a new facet in the co-evolutionary processes that shape insect-plant interactions at a molecular level, whereby herbivore-derived elicitors denoted as Herbivore-Associated Molecular Patterns (HAMPs)^21,22 mediate the reprogramming of HPL-pathway genes. Among several intriguing questions arising from these observations are: the chemical nature of insect OS component responsible for suppression of HPL pathway, and mechanisms by which signals are sensed and relayed into downstream signaling cascades involved in constitutive deactivation of HPL-defense responsive pathways.

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.