FOUR SERINE PROTEASE cDNAS FROM THE MIDGUT OF Plutella xylostella AND THEIR PROTEINASE ACTIVITY ARE INFLUENCED BY THE ENDOPARASITOID, Cotesia vestalis

Abstract

Serine proteinases, which include trypsins and chymotrypsins, play numerous roles in lepidopteran larvae, such as digestion, zymogen activation, and immune defense. Studies of lepidopteran serine proteinases could increase understanding of their feeding preference (polyphagous and monophagous) and facilitate identification of protease inhibitors, which can be engineered for pest management. In this paper, four full-length cDNAs encoding one chymotrypsin and three trypsins were cloned from larval midguts of the diamondback moth, Plutella xylostella. Real-time quantitative polymerase chain reaction (PCR) analysis showed all four serine protease genes were downregulated after P. xylostella was parasitized by the parasitoid wasp Cotesia vestalis. Trypsin and chymotrypsin enzymatic activities within the midgut of nonparasitized and C. vestalis-parasitized P. xylostella larvae were examined using N-a-benzoyl-arg p-nitroanilide and N-succinyl-ala-ala-pro-phe p-nitroanilide as substrates. Trypsin and chymotrypsin activities were decreased in parasitized larvae as compared to untreated larvae, with the effect being more pronounced over time. Chymotrypsin activity in particular exhibited a significant decrease in activity. The correlation of decreased enzymatic activity and transcript abundance suggests parasitization induced downregulation of serine proteinase gene transcripts.