The Gut Transcriptome Of Hessian Fly, Mayetiola Destructor (Say), And The Gene Expression Of Differential Responses Of Wheat Inhibitor-Like Genes To The Insect Attacks

The Hessian fly, Mayetiola destructor (Say), is a destructive pest of wheat and is widely distributed throughout most wheat-growing areas of the world. Host plant resistance has been considered the most effective and economical means of controlling this pest in wheat. However, the widespread use of resistant cultivars has placed a strong selection pressure on Hessian fly populations, which has resulted in the evolution of new genotypes of the fly that can overcome previously resistant wheat. Since the gut is the primary site for food digestion, nutrient absorption, toxic degradation and plant resistance. Thus, identification and characterization of genes expressed in the Hessian fly larval gut will not only increase our understanding of insect gut physiology, but provide novel and specific targets for the development of strategies for pest management. On the other hand, the wheat-Hessian fly system follows a typical gene-for-gene interaction, and provides a good model for studying the scheme of attack and counter-attack between insects and plants. In the paper, the first global analysis of gut transcripts from a gall midge and systematic analysis of the expression patterns of inhibitor-like genes in wheat-Hessian fly system were conducted by means of methods and techniques of Expressed sequenced tag (ESTs), transcriptome, microarray and Real-Time PCR. The main results were as follows.1. More than 10 000 ESTs were assembled into 2 024 clusters (contigs and singletons). BLASTx identified 1 216 clusters with similarity to GenBank sequences. Among them, 809 clusters coded either for proteins with similarity to functionally known proteins or for small secretory proteins (SSP) (<250 amino acids). These clusters were grouped into nine categories: I. Proteins involved in metabolism, II. Structural proteins, III. Regulators, IV. Transporters, V. Proteins involved in protein synthesis and folding, VI. Digestive enzymes, VII. Detoxification enzymes, VIII. SSP, and IX. Others. The first five categories represent genes participating in house-keeping functions. The other three categories, including digestive enzymes, detoxification enzymes, and SSP, are characteristic of gut functions.2. There were transcripts coding for diverse putative digestive proteases, which is different from other plant-sucking insects such as aphids. The digestive enzymes included clusters coding for 10 trypsins (six represented novel genes, named MDP6A, MDP6B, MDP7A, MDP8C, MDP9A and MDP10A), seven chymotrypsins (four represented novel genes, named MDP11A, MDP12B, MDP13A and MDP14B), two cysteine proteases (novel genes), one aspartic protease (novel genes), one endo-oligopeptidase (novel genes), three aminopeptidases (novel genes), 10 carboxypeptidases (one novel gene), and twoα-amylases (novel genes).3. Detoxification enzymes included clusters coding for 13 cytochrome P450s (novel genes), three glutathione S-transferases (GSTs) (one novel gene), 3 peroxidases (one gene), three ferritins (novel genes), one catalase (novel gene), two peroxiredoxins (novel genes), and several other enzymes. The existence of transcripts coding for diverse antioxidant enzymes may be the molecular basis for Hessian fly to overcome toxic oxidants and secondary metabolites resulted from basal and induced defenses of host plants.4. The category of SSP contained 111 clusters, 22 of which coded for proteins structurally similar to protease inhibitors (seven novel genes), ribonucleases (two novel genes), cuticle proteins (four novel genes), and cellular regulators, whereas the other clusters coded for unknown proteins. The unique feature of Hessian fly larval gut transcriptome was the presence of a large number of transcripts coding for SSP, which represented nearly 25% of all the ESTs. Most (63%) of the putative SSP were unique to Hessian fly or gall midges. This strongly suggests that they probably perform functions characteristic of this insect, such as serving as effectors for the inhibition of host growth or in establishing nutritive tissue at the feeding site.5. Four groups of inhibitor-like genes encoding proteins with diverse structures were identified from wheat. The majorities of these genes were up-regulated by avirulent Hessian fly larvae during incompatible interactions, and were down-regulated by virulent larvae during compatible interactions. The upregulation during incompatible interactions and downregulation during compatible interactions resulted in 4 to 30 fold differences between the expression levels in resistant plants and those in susceptible plants. The increased expression of inhibitor-like genes during incompatible interactions suggested that these genes are part of defense mechanisms in wheat against Hessian fly attacks, whereas the downregulation of these genes during compatible interactions suggested that virulent larvae can suppress plant defenses. Both the upregulation of the inhibitor-like genes during incompatible interactions by avirulent larvae and the downregulation during compatible interactions by virulent larvae were through mechanisms that were independent of the wound response pathway.