Abstract
BackgroundTrehalase, an enzyme that hydrolyzes trehalose to yield two glucose molecules, plays a pivotal role in various physiological processes. In recent years, trehalase proteins have been purified from several insect species and are divided into soluble (Tre-1) and membrane-bound (Tre-2) trehalases. However, no functions of the two trehalases in chitin biosynthesis in insects have yet been reported.Principal FindingsThe membrane-bound trehalase of Spodoptera exigua (SeTre-2) was characterized in our laboratory previously. In this study, we cloned the soluble trehalase gene (SeTre-1) and investigated the tissue distribution and developmental expression pattern of the two trehalase genes. SeTre-1 was expressed highly in cuticle and Malpighian tubules, while SeTre-2 was expressed in tracheae and fat body. In the midgut, the two trehalase genes were expressed in different locations. Additionally, the expression profiles of both trehalase mRNAs and their enzyme activities suggest that they may play different roles in chitin biosynthesis. The RNA interference (RNAi) of either SeTre-1 or SeTre-2 was gene-specific and effective, with efficiency rates up to 83% at 72 h post injection. After RNAi of SeTre-1 and SeTre-2, significant higher mortality rates were observed during the larva-pupa stage and pupa-adult stage, and the lethal phenotypes were classified and analyzed. Additionally, the change trends of concentration of trehalose and glucose appeared reciprocally in RNAi-mutants. Moreover, knockdown of SeTre-1 gene largely inhibited the expression of chitin synthase gene A (CHSA) and reduced the chitin content in the cuticle to two-thirds relative to the control insects. The chitin synthase gene B (CHSB) expression, however, was inhibited more by the injection of dsRNA for SeTre-2, and the chitin content in the midgut decreased by about 25%.Conclusions SeTre-1 plays a major role in CHSA expression and chitin synthesis in the cuticle, and SeTre-2 has an important role in CHSB expression and chitin synthesis in the midgut.
Highlights
Trehalase (a-glucoside-1-glucohydrolase, EC 3.2.1.28) is an enzyme that hydrolyzes trehalose to yield two glucose molecules, and it is present in almost all tissues in different forms
SeTre-1 plays a major role in chitin synthase gene A (CHSA) expression and chitin synthesis in the cuticle, and SeTre-2 has an important role in chitin synthase gene B (CHSB) expression and chitin synthesis in the midgut
The chitin biosynthesis pathway starts with trehalose[5], and the first enzyme involved in the pathway is trehalase, and the last one is chitin synthase
Summary
Trehalase (a-glucoside-1-glucohydrolase, EC 3.2.1.28) is an enzyme that hydrolyzes trehalose to yield two glucose molecules, and it is present in almost all tissues in different forms. Trehalase plays a pivotal role in various physiological processes, including flight metabolism[1], chitin synthesis during molting [2], and cold tolerance[3]. In insects, all these function of trehalase are achieved through the hydrolysis of trehalose (a-D-glucopyranosyl-a-Dglucopyranoside), the principal hemolymph sugar in insects that acts as an indispensable substrate for energy production and macromolecular biosynthesis[4]. An enzyme that hydrolyzes trehalose to yield two glucose molecules, plays a pivotal role in various physiological processes. Trehalase proteins have been purified from several insect species and are divided into soluble (Tre-1) and membrane-bound (Tre-2) trehalases. No functions of the two trehalases in chitin biosynthesis in insects have yet been reported
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