Detection and functional characterization of sigma class GST in Phlebotomus argentipes and its role in stress tolerance and DDT resistance

Abstract

AbstractSeveral Glutathione S-transferases (GSTs) enzymes, in insects, have previously been implicated in resistance developed against DDT and other insecticides. The GST enzyme particularly sigma class have important physiological role in detoxification of lipid peroxidation by-products in insects.Phlebotomus argentipeshas been intensely exposed to DDT over years due to Indoor Residual Spray (IRS) programme for Kala-azar elimination in Bihar, India. However, inP. argentipes, role of GSTs in DDT resistance have not been elucidated. Here, sigma class GST ofP. argentipes(Parg-GSTσ) was successfully cloned, expressed and purified by affinity chromatography. The recombinant Parg-GSTσ was found to be highly active towards cumene hydroperoxide and 4-HNE having specific activity 92.47 & 203.92 µM/min/mg of protein, respectively and exhibited low activity towards universal substrate CDNBi.e., 8.75 µM/min/mg of protein. RT-PCR and immunoblot analysis showed at least 2 and 1.8 fold overexpression of Parg-GSTσ in the single exposed and non exposed DDT resistantP. argentipesas compared to susceptible, implicating Parg-GSTσ also involved in DDT resistance probably by imparting enhanced stress tolerance. The DDT, H2O2and temperature induction assays demonstrated stress-dependent induction of Parg-GSTσ expression indicating its important role in oxidative stress redressal.