Expression analysis and RNA interference of BmCarE-10 gene from Bombyx mori

Mol Biol Rep. 2014 Mar;41(3):1607-16. doi: 10.1007/s11033-013-3007-3. Epub 2014 Jan 11.

Abstract

Carboxylesterase (CarE) is a multifunctional superfamily, and it plays important roles in xenobiotic detoxification, pheromone degradation, neurogenesis and regulating development. In this research, firstly, we measured the rutin-induced transcriptional level of BmCarE-10 gene by using real-time quantitative RT-PCR method, and dual spike-in strategy. Several possible physiological functions were certified preliminarily by RNAi experiments in silkworm. Promoter truncation analysis using a dual-luciferase reporter assay in Bombyx mori ovary cells (BmN) showed that the region -705 to -625 for BmCarE-10 gene was essential for basal and rutin-induced transcriptional activity. Sequence analysis of this region revealed several potential transcriptional regulatory elements such as Croc and Dfd. The activities of the BmCarE-10 promoter in various tissues of silkworm were also measured by firefly luciferase activity and normalized by the Renilla luciferase activity. Results showed that the activity of the BmCarE-10 promoter were highest in the Malpighian tubule, followed by fat body, silk gland, midgut, epidermis, and hemocyte. The essential region for basal and rutin-induced transcriptional activity was also -894 to -502 in Malpighian tubule and fat body of silkworm. The potential core promoters of BmCarE-10 gene in B. mori are reported for the first time in this research. Further identification of cis- and trans-elements and their role in upregulation of BmCarE-10 gene may be useful for elucidating the contribution of CarE protein to the response mechanism to rutin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bombyx / genetics*
  • Bombyx / metabolism
  • Carboxylesterase / biosynthesis*
  • Carboxylesterase / genetics
  • Cloning, Molecular
  • Female
  • Gene Expression Regulation, Enzymologic / drug effects
  • Gene Expression Regulation, Enzymologic / genetics
  • Larva
  • Ovary / cytology
  • Ovary / metabolism*
  • Promoter Regions, Genetic
  • RNA Interference*
  • Rutin / pharmacology

Substances

  • Rutin
  • Carboxylesterase