Abstract: 【Aims】 Honey bees are of the holometabolous insects, whose brood is typically stenothermic. In order to understand the key genes involved in the sensitivity of the Italian honey bee, Apis mellifera ligustica to cold stress, we conducted a trend analysis of differentially expressed genes (DEGs) of cold-sensitive prepupae in response to cold stress. 【Methods】 The 3 d-old post-capped prepupae were exposed to cold temperature (20℃) for 18 (T18) and 36 h (T36), respectively, with those unexposed to cold stress used as the control (CK), and then subjected to transcriptomic sequencing based on Illumina HiSeq^TM. Short Time-series Expression Miner (STEM) software was applied to compare the shared DEGs among the three datasets. In addition, GO and KEGG pathway enrichment analyses were performed for DEGs with up- and down-regulation profiles. The expression profiles of five randomly selected DEGs were verified by RT-qPCR. 【Results】 Trend analysis showed that 1 062 DEGs shared by T18 vs CK and T36 vs CK were clustered into three significant profiles including two up-regulation profiles (Profile 6 covering 539 DEGs and Profile 7 covering 271 DEGs) and one down-regulation profile (Profile 1 covering 183 DEGs). GO and KEGG pathway enrichment analyses showed that insulin-like peptide gene (ILP) and forkhead box protein O gene (FoxO) were up-regulated in both FoxO signaling and longevity regulating pathways within Profile 6, suggesting that ecdysone signaling responds to cold stress during honey bee pupation. The CREB-binding protein gene (CBP) in Profile 7 was up-regulated, and CYP450 306a1 (phm) and WNT1 in Profile 1 were down-regulated, suggesting that these genes regulate cell development and ecdysone biosynthesis in response to low temperature. Besides, the expression profiles of the five selected genes based on RT-qPCR analysis were consistent with those based on the transcriptome. 【Conclusion】 The trend analysis of DEGs reveals that FoxO regulated by insulin signaling and ecdysone signaling in honey bees may act as a key gene that represses honey bee pupation in response to cold stress. Our data lay a foundation for exploring the molecular regulation mechanisms underlying metamorphosis in response to cold stress in honey bees.

Key words: Apis mellifera ligustica, pupation, low temperature stress, transcriptomics, ecdysone