Identification of an aromatic aldehyde synthase involved in indole-3-acetic acid biosynthesis in the galling sawfly (Pontania sp.) and screening of an inhibitor.

Indole-3-acetic acid (IAA), a phytohormone auxin, may be involved in insect gall induction. We previously proposed that the IAA biosynthetic pathway is Trp → indole-3-acetaldoxime → indole-3-acetaldehyde (IAAld) → IAA or Trp → IAAld → IAA. In this study, we surveyed galling sawfly enzymes responsible for the rate-limiting steps using a heterologous protein expression system and identified PonAAS2, an aromatic aldehyde synthase, that catalyzed the conversion of Trp to IAAld. The PonAAS2 gene was highly expressed in early- and mid-stage larvae that contained high concentrations of IAA, but the expression level was almost negligible in larvae that had escaped from their gall in autumn and contained very low concentrations of IAA. An inhibitor of PonAAS2, obtained by screening a chemical library, inhibited IAA production in sawfly enzyme solution by 80%, suggesting the important role of this enzyme in IAA biosynthesis in sawfly.

Identification of an aldehyde oxidase involved in indole-3-acetic acid synthesis in Bombyx mori silk gland.

Auxin is thought to be an important factor in the induction of galls by galling insects. We have previously shown that both galling and nongalling insects synthesize indole-3-acetic acid (IAA) from tryptophan (Trp) via two intermediates, indole-3-acetaldoxime (IAOx) and indole-3-acetaldehyde (IAAld). In this study, we isolated an enzyme that catalyzes the last step "IAAld → IAA" from a silk-gland extract of Bombyx mori. The enzyme, designated "BmIAO1", contains two 2Fe-2S iron-sulfur-cluster-binding domains, an FAD-binding domain, and a molybdopterin-binding domain, which are conserved in aldehyde oxidases. BmIAO1 causes the nonenzymatic conversion of Trp to IAAld and the enzymatic conversion of IAOx to IAA, suggesting that BmIAO1 alone is responsible for IAA production in B. mori. However, a detailed comparison of pure BmIAO1 and the crude silk-gland extract suggested the presence of other enzymes involved in IAA production from Trp. Abbreviations: BA: benzoic acid; CE: collision energy; CXP: collision cell exit potential; DP: declustering potential; IAA: indole-3-acetic acid; IBI1: IAA biosynthetic inhibitor-1; IAAld: indole-3-acetaldehyde; ICA: indole-3-carboxylic acid; IAOx: indole-3-acetaldoxime; IEtOH: indole-3-ethanol; LC-MS/MS: liquid chromatography-tandem mass spectrometry; Trp: tryptophan.

Novel tryptophan metabolic pathways in auxin biosynthesis in silkworm.

In the course of our study of the biosynthetic pathway of auxin, a class of phytohormones, in insects, we proposed the biosynthetic pathway tryptophan (Trp)→indole-3-acetaldoxime (IAOx)→indole-3-acetadehyde (IAAld)→indole-3-acetic acid (IAA). In this study, we identified two branches in the metabolic pathways in the silkworm, possibly affecting the efficiency of IAA production: Trp→indole-3-pyruvic acid→indole-3-lactic acid and IAAld→indole-3-ethanol. We also determined the apparent conversion activities (2.05×10-7UmL-1 for Trp→IAA, 1.30×10-5UmL-1 for IAOx→IAA, and 3.91×10-1UmL-1 for IAAld→IAA), which explain why IAOx and IAAld are barely detectable as either endogenous compounds or metabolites of their precursors. The failure to detect IAAld, even in the presence of an inhibitor of the conversion IAAld→IAA, is explained by a switch in the conversion from IAAld→IAA to IAAld→IEtOH.

Transcriptomic characterization of gall tissue of Japanese elm tree (Ulmus davidiana var. japonica) induced by the aphid Tetraneura nigriabdominalis.

Insect galls are abnormal plant tissues induced by parasitic insect(s) for use as their habitat. In previous work, we suggested that gall tissues induced by the aphid Tetraneura nigriabdominalis on Japanese elm trees are less responsive than leaf tissues to jasmonic acid (JA), which is involved in the production of volatile organic compounds as a typical defensive reaction of plants against attack by insect pests. A comprehensive analysis of gene expression by RNA sequencing indicated that the number of JA responsive genes was markedly lower in gall tissues than in leaf tissues. This suggests that gall tissues are mostly defective in JA signaling, although JA signaling is not entirely compromised in gall tissue. Gene ontology analysis sheds light on some stress-related unigenes with higher expression levels in gall tissues, suggesting that host plants sense aphids as a biotic stress but are defective in the JA-mediated defense response in gall tissues.

Adaptive significance of gall formation for a gall-inducing aphids on Japanese elm trees.

Insect galls are abnormal plant tissues induced by external stimuli from parasitizing insects. It has been suggested that the stimuli include phytohormones such as auxin and cytokinins produced by the insects. In our study on the role of hormones in gall induction by the aphid Tetraneura nigriabdominalis, it was found that feedback regulation related to auxin and cytokinin activity is absent in gall tissues, even though the aphids contain higher concentrations of those phytohormones than do plant tissues. Moreover, jasmonic acid signaling appears to be compromised in gall tissue, and consequently, the production of volatile organic compounds, which are a typical defense response of host plants to herbivory, is diminished. These findings suggest that these traits of the gall tissue benefit aphids, because the gall tissue is highly sensitive to auxin and cytokinin, which induce and maintain it. The induced defenses against aphid feeding are also compromised. The abnormal responsiveness to phytohormones is regarded as a new type of extended phenotype of gall-inducing insects.