Effects of different temperature regimes on the expression of the trehalose-6-phosphate synthase gene and haemolymph sugar content in Gampsocleis gratiosa
Author of the article:QIAN Lei-Yang** KOU Xiao-Yan DONG Ze-Hua CHANG Yan-Lin ZHOU Zhi-Jun***
Author's Workplace：Key Laboratory of Invertebrate Systematics and Application of Hebei Province, College of Life Science, Hebei University, Baoding 071002, China
Key Words：TPS gene, trehalose, total sugar, temperature, real-time fluorescent quantitative PCR
[Objectives] To ascertain the effects of rapid vs gradual change in temperature on trehalose-6-phosphate synthase (TPS) gene expression levels, and trehalose, and total sugar, content, in the haemolymph of Gampsocleis gratiosa. [Methods] Based on the G. gratiosa transcriptome sequencing data set, the full cDNA of the G. gratiosa TPS gene was cloned. Rapid and gradual change in temperature treatments were applied to G. gratiosa. The body temperature of G. gratiosa was measured with a thermocouple thermometer, and the environmental temperature closest to body temperature was regarded as the optimal temperature. TPS gene expression was detected using real-time fluorescent quantitative PCR. Sulfuric acid-anthrone colorimetry was used to measure trehalose, and total sugar, content (the control was the normal content at 25℃). [Results] The cDNA was cloned and designated as GgTPS (GenBank accession no. KU578006). The full-length cDNA of GgTPS is 3 225 bp with 5′- and 3′-untranslated regions of 153 bp and 642 bp, respectively. It contains an open reading frame of 2 430 nucleotides encoding a protein of 809 amino acids residues with a predicted molecular mass of approximately 91.35 ku and isoelectric point of 6.28. TPS expression levels first increased, then decreased, peaking at 15℃, which was in marked contrast to that measured at 25℃ during the rapid cooling phase. Trehalose content remained constant level before increasing to a high level at 0℃. Total sugar content increased gradually, reaching a maximal level at 0℃. During the rapid heating stage, TPS had a down-up-down expression profile, peaking at 40℃. During the gradual cooling stage, TPS expression first increased, then decreased, and was highly expressed at 15℃ and 20℃. Trehalose content first decreased, then increased, and was significantly higher at 0℃. Total sugar content was markedly different at 0℃, 10℃ and 20℃ to that measured at 25℃. During gradual heating, TPS expression first declined, then increased, and was significantly lower at 30℃ and 45℃. Trehalose content was significantly lower at 40℃ than at 25℃. Total sugar content first increased, then declined, and was significantly higher at 40℃ and 45℃ than at 25℃. Comparing rapid vs gradual change in temperature, TPS expression differed significantly at 15℃ and 40℃, and trehalose content was markedly different at 40℃ and 45℃. Total sugar content was markedly different at 10℃ and 20℃, and body temperature was also markedly different at 30℃. [Conclusion] Temperature acclimatization improves the ability of insects to resist stress. The TPS expression levels and trehalose content of insects are closely related to ambient temperature, as well as the duration of temperature acclimatization. Patterns of TPS expression and trehalose accumulation in G. gratiosa kept at low temperature differ from those at high temperature.