Abstract
The Omega-class of GSTs (GSTOs) is a class of cytosolic GSTs that have specific structural and functional characteristics that differ from those of other GST groups. In this study, we demonstrated the involvement of the GSTO1 gene from A. cerana cerana in the oxidative stress response and further investigated the effects of three cysteine residues of GSTO1 protein on this response. Real-time quantitative PCR (qPCR) showed that AccGSTO1 was highly expressed in larvae and foragers, primarily in the midgut, epidermis, and flight muscles. The AccGSTO1 mRNA was significantly induced by cold and heat at 1 h and 3 h. The TBA (2-Thiobarbituric acid) method indicated that cold or heat resulted in MDA accumulation, but silencing of AccGSTO1 by RNAi in honeybees increased the concentration of MDA. RNAi also increased the temperature sensitivity of honeybees and markedly reduced their survival. Disc diffusion assay indicated that overexpression of AccGSTO1 in E. coli caused the resistance to long-term oxidative stress. Furthermore, AccGSTO1 was active in an in vitro DNA protection assay. Mutations in Cys-28, Cys-70, and Cys-124 affected the catalytic activity and antioxidant activity of AccGSTO1. The predicted three-dimensional structure of AccGSTO1 was also influenced by the replacement of these cysteine residues. These findings suggest that AccGSTO1 plays a protective role in the response to oxidative stress.
Highlights
Reactive oxygen species (ROS), such as oxygen radical superoxide (O2–) and hydroxyl (OH–), are constantly generated during aerobic metabolism
Compared to Bombus terrestris and Nasonia vitripennis, GSTO1 from A. cerana cerana was placed closest to Apis mellifera and Apis florea
There are five introns in both AccGSTO genes, the first intron of AccGSTO1 is located in the 59 untranslated region (UTR) (Fig. 1C)
Summary
Reactive oxygen species (ROS), such as oxygen radical superoxide (O2–) and hydroxyl (OH–), are constantly generated during aerobic metabolism. Three Omega GST genes have been found in the human genome [18], two GSTO genes have been found in mice and rats [11], and two GSTO genes have been identified in honeybees [8] These GSTOs display distinct genetic organizations, crystal structures, substrate specificities, and catalytic activities [19]. GSTO1 exhibits a canonical cytosolic GST fold, it has unique Nterminal and C-terminal extensions that are not observed in the other GST classes. These two domains of GSTO1 interact to form a distinct structural unit that is crucial for substrate specificity [20]. Our findings provide valuable insight into the function of GSTO1 in the oxidative stress response
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