Molecular Characterization Of Small Heat Shock Proteins And Their Responses Under Cadmium Stress In Oxya Chinensis

Small heat shock proteins (sHSPs) are the most diverse proteins in terms of structure and function among the various stress protein families. They display chaperone-like activity and help unfolding proteins maintain their correct states. sHSPs have been implicated in many physiological processes and play important roles in the response to various stresses. In the study, molecular characterization of small heat shock proteins and their responses under cadmium stress in Oxya chinensis was analyzed, the results may provide basic data for further study of OcsHSPs.1. In this study, the full-length sequences of six sHSP genes were obtained from the Oxya chinensis transcriptome database. The sequences were confirmed using the specific primers OcHSP19.1,19.8,20.4,20.7,21.1 and 23.8 and their molecular weights. The deduced amino acid sequences of the six OcsHSPs contain a typical α-crystallin domain, which consists of approximately 100 amino acid residues and five β-strands. The phylogenetic analysis suggested that OcHSP23.8 was orthologous to the sHSPs of other species and that OcHSP19.1,20.4,20.7 and 21.1 were species-specific, whereas OcHSP19.8 did not cluster closely to Orthoptera but was placed on the basal end of the cluster. The cloning and sequence analysis of OcsHSPs may lay a foundation for the research of their gene expression patterns and functions.2. Developmental stage-dependent and tissue-specific expression patterns were evaluated using real time quantitative PCR (RT-qPCR). The six OcsHSP genes were expressed in all developmental stages. The expression patterns of the six genes showed clear tissue specificity, the highest expression levels for OcHSP19.1,20.4,20.7 and 21.1 were in the ovary and/or testis, the highest expression levels of OcHSP19.8 and 23.8 were in the muscles. These sHSP genes that are expressed in special tissues may play an important role in maintaining the normal functions of cells and tissues. The testis and ovary are the primary reproductive organs, these results suggest that these sHSPs may participate in reproduction and development. Developmental stage-dependent expression pattern reported that they are involved in the regulation of insect development. The tissue and development expression of OcsHSP genes will help to further study on their function.3. Small heat shock proteins (sHSPs) can be induced by environmental stresses. In the present study, we first analyzed the mRNA expression patterns of the six OcsHSP genes in the fifth-instar nymphs of Oxya chinensis exposed to different temperatures using real-time quantitative PCR. The results showed the expression levels of OcHSP 19.1, OcHSP19.8, OcHSP20.4 and OcHSP20.7 had no obvious changes at 27,32 and 37℃ of different times compared with the controls, but their expression levels significantly increased at 42℃, especially at 42℃ of 2 h. The expression levels of OcHSP21.1 and OcHSP23.8 had no obvious changes at different temperatures compared with the controls. The results show that the six OcsHSPs gene have different response to temperature changes.4. Later, we analyzed the mRNA expression patterns of the six OcsHSP genes in the whole body, testis and ovary of the fifth-instar nymphs of Oxya chinensis exposed to cadmium (Cd) using real-time quantitative PCR. The results indicate that Cd2+ can induce the six genes. However, various response patterns were observed among these genes under Cd2+ stress conditions. In the whole body, OcHSP 19.1,19.8,20.4 and 20.7 were highly induced by 0.48μg/μL Cd2+ at 24 h; OcHSP23.8 was significantly up-regulated by 0.48 μg/μL Cd2+ at 6 h; for OcHSP21.1, the highest expression levels were found after treatment with 0.16μg/μL Cd2+ for 24 h,0.32 μg/μL Cd2+for 36 h, and 0.48μg/μL Cd2+ for 12 h. In the testis,0.48μg/μL Cd2+ induced the expression of OcHSP19.1 and OcHSP 19.8 genes in different treatment times, both reached the highest level at 36 h. In the ovary, when treated by 0.32 μg/μL Cd2+, the expression levels of OcHSP 19.1 and OcHSP21.1 gene both reached the highest level at 36 h, the expression levels of OcHSP20.7 gene first increased then declined significantly. The expression levels of OcHSP21.1 gene in the testis and OcHSP20.4 gene in the ovary had no significant changes compared with the controls. The results showed that the expression patterns of the five OcsHSP genes related to the Cd concentrations and treatment times. The various response patterns of the different genes under Cd2+ stress suggest that there are differences in the biological functions of these genes.In this study, the full-length sequences of six sHSP genes were cloned from the Oxya chinensis, the secondary structure and phylogenetic analysis was conducted. Developmental stage-dependent and tissue-specific expression patterns were evaluated using real time quantitative PCR (RT-qPCR), then we analyzed the mRNA expression patterns of the six OcsHSP genes in the fifth-instar nymphs of Oxya chinensis exposed to different temperatures and cadmium. To sum up, this paper systemicly analysed OcsHSPs at the genetic level, these provide basic data for further researching the physiological and biochemical function of them, in order to laying a theoretical foundation for OcsHSPs as molecular markers of acute cadmium pollution.