Acta Entomologica Sinica ›› 2019, Vol. 62 ›› Issue (5): 535-546.doi: 10.16380/j.kcxb.2019.05.002

• RESEARCH PAPERS • Previous Articles     Next Articles

Cloning of two clip-domain serine protease genes and their expression in response to exogenous hormone and immune challenge in Lasioderma serricorne (Coleoptera: Anobiidae)(In English)

CHEN Chun-Xu1,2, XU Kang-Kang1,2, YANG Hong1,3,*, ZHAO Feng1, YAN Yi1,2, HU Da-Ming4, YANG Wen-Jia1,2   

  1. (1. Provincial Key Laboratory for Agricultural Pest Management of Mountainous Regions, Institute of Entomology, Guizhou University, Guiyang 550025, China; 2. Guizhou Provincial Key Laboratory for Rare Animal and Economic Insect of the Mountainous Region, College of Biology and Environmental Engineering, Guiyang University, Guiyang 550005, China; 3. College of Tobacco Science, Guizhou University, Guiyang 550025, China; 4. China Tobacco Guizhou Industrial Co., LTD, Guiyang 550005, China)
  • Online:2019-05-20 Published:2019-05-16


【Aim】 As essential components of extracellular signaling cascades, clip-domain serine proteases (CLIPs) play important roles in insect development and innate immunity. This study aims to clone two CLIP genes from the cigarette beetle, Lasioderma serricorne, to clarify their developmental stage- and tissue-specific expression profiles, and to analyze their expression changes in response to exogenous hormone 20-hydroxyecdysone (20E) and immune challenge, so as to lay the foundation for further understanding the physiological functions of CLIPs in L. serricorne. 【Methods】 The full-length cDNAs of two CLIP-encoding genes (LsCLIP1 and LsCLIP2) were cloned using RT-PCR, and the deduced protein structure was predicted using different bioinformatics software, and the phylogenetic tree of insect CLIPs was constructed using the MEGA 6.06 program. The mRNA expression profiles of these two genes in different developmental stages [early larva (<24 h post egg hatching), late larva (older than the 4th instar), pupa (>48 h post pupation), early adult (<24 h post eclosion), and late adult (7 d post eclosion)] and different tissues (cuticle, fat body, gut, and carcass) of the 5th instar larvae, and the 4th instar larvae of L. serricorne injected with 20E (120 ng/larva) and peptidoglycan (0.2 μL) from Escherichia coli or Staphylococcus aureus were detected by quantitative real-time PCR (qPCR). 【Results】 The full-length cDNA sequences of LsCLIP1 and LsCLIP2 were cloned from L. serricorne, and both genes have an open reading frame of 1 194 bp encoding 397 amino acid residues. The predicted CLIP1 and CLIP2 proteins both contain a typical clip domain and a trypsin-like serine protease domain. Phylogenetic analyses revealed that CLIP1 and CLIP2 belong to subfamily C CLIPs. The qPCR analyses confirmed that both LsCLIP1 and LsCLIP2 were expressed throughout the developmental stages and in all the assayed larval tissues of L. serricorne, with particularly high expression level in the pupal stage and the cuticle, respectively. Expression of both genes was significantly upregulated in larvae exposed to 20E and following challenge with peptidoglycan from E. coli or S. aureus. 【Conclusion】 It is suggested that LsCLIP1 and LsCLIP2 might be involved in insect molting and innate immune response. These results provide valuable information for further study on the regulation of CLIPs in insects.

Key words: Lasioderma serricorne, clip-domain serine proteases, 20-hydroxyecdysone, immune challenge, expression patterns