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Identification and characterization of an Apis cerana cerana Delta class glutathione S-transferase gene (AccGSTD) in response to thermal stress

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Abstract

Glutathione S-transferases (GSTs) are members of a multifunctional enzyme super family that plays a pivotal role in both insecticide resistance and protection against oxidative stress. In this study, we identified a single-copy gene, AccGSTD, as being a Delta class GST in the Chinese honey bee (Apis cerana cerana). A predicted antioxidant response element, CREB, was found in the 1,492-bp 5′-flanking region, suggesting that AccGSTD may be involved in oxidative stress response pathways. Real-time PCR and immunolocalization studies demonstrated that AccGSTD exhibited both developmental- and tissue-specific expression patterns. During development, AccGSTD transcript was increased in adults. The AccGSTD expression level was the highest in the honey bee brain. Thermal stress experiments demonstrated that AccGSTD could be significantly upregulated by temperature changes in a time-dependent manner. It is hypothesized that high expression levels might be due to the increased levels of oxidative stress caused by the temperature challenges. Additionally, functional assays of the recombinant AccGSTD protein revealed that AccGSTD has the capability to protect DNA from oxidative damage. Taken together, these data suggest that AccGSTD may be responsible for antioxidant defense in adult honey bees.

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Acknowledgments

This work was funded by the China Agriculture Research System (No. CARS-45), Agro-scientific Research in the Public Interest (No. 200903006), and the National Natural Science Foundation (No. 31172275) in China.

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Authors and Affiliations

  1. State Key Laboratory of Crop Biology, College of Life Sciences, Shandong Agricultural University, Tai’an, Shandong, 271018, People’s Republic of China

    Huiru Yan, Haihong Jia, Xiuling Wang, Hongru Gao & Xingqi Guo

  2. College of Animal Science and Technology, Shandong Agricultural University, Tai’an, Shandong, 271018, People’s Republic of China

    Baohua Xu

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  1. Huiru Yan
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  2. Haihong Jia
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  3. Xiuling Wang
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  4. Hongru Gao
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  5. Xingqi Guo
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Correspondence to Xingqi Guo or Baohua Xu.

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Communicated by: Sven Thatje

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Supplementary Fig. 1

Standard curves of AccGSTZ1 and AccGSTD. a Standard curve of AccGSTZ1 gene from the amplification of six ten-fold serial dilutions of plasmid fused by AccGSTZ1. b Standard curve of AccGSTD gene from the amplification of six ten-fold serial dilutions of the same plasmid fused by AccGSTD. Correlation coefficient and slope values are indicated. The calculated threshold cycle values were plotted versus the log of each starting quantity. (JPEG 24 kb)

High resolution image (TIFF 3640 kb)

Supplementary Fig. 2

Determination of anti-AccGSTD specificity using immunoblot analysis (a) and ponceau staining (b). Ponceau staining to verify similar amounts of protein was loaded into each lane. Lane M: pre-stained protein molecular weight marker; lane 1: uninduced recombinant AccGSTD protein; lanes 2: induced recombinant AccGSTD protein; lane 3: purified recombinant AccGSTD protein. (JPEG 7 kb)

High resolution image (TIFF 6764 kb)

Supplementary Table 1

Oligonucleotide primers used in this study (DOCX 15 kb)

Supplementary Table 2

PCR amplification conditions in this study (DOCX 14 kb)

Supplementary Table 3

Estimation of copy number of AccGSTD gene in Apis cerana cerana (DOCX 13 kb)

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Yan, H., Jia, H., Wang, X. et al. Identification and characterization of an Apis cerana cerana Delta class glutathione S-transferase gene (AccGSTD) in response to thermal stress. Naturwissenschaften 100, 153–163 (2013). https://doi.org/10.1007/s00114-012-1006-1

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  • DOI: https://doi.org/10.1007/s00114-012-1006-1

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