Induction of soluble AChE expression via alternative splicing by chemical stress in Drosophila melanogaster

Abstract

Various molecular forms of acetylcholinesterase (AChE) have been characterized in insects. Post-translational modification is known to be a major mechanism for the molecular diversity of insect AChE. However, multiple forms of Drosophila melanogaster AChE (DmAChE) were recently suggested to be generated via alternative splicing (Kim and Lee, 2013). To confirm alternative splicing as the mechanism for generating the soluble form of DmAChE, we generated a transgenic fly strain carrying the cDNA of DmAChE gene (Dm_ace) that predominantly expressed a single transcript variant encoding the membrane-anchored dimer. 3′ RACE (rapid amplification of cDNA ends) and western blotting were performed to compare Dm_ace transcript variants and DmAChE forms between wild-type and transgenic strains. Various Dm_ace transcripts and DmAChE molecular forms were observed in wild-type flies, whereas the transgenic fly predominantly expressed Dm_ace transcript variant encoding the membrane-anchored dimer. This supports alternative splicing as the major determinant in the generation of multiple forms of DmAChE. In addition, treatment with DDVP as a chemical stress induced the expression of the Dm_ace splice variant without the glycosylphosphatidylinositol anchor site in a dose-dependent manner and, accordingly, the soluble form of DmAChE in wild-type flies. In contrast, little soluble DmAChE was expressed in the transgenic fly upon exposure to DDVP. DDVP bioassays revealed that transgenic flies, which were unable to express a sufficient amount of soluble monomeric DmAChE, were more sensitive to DDVP compared to wild-type flies, suggesting that the soluble monomer may exert non-neuronal functions, such as chemical defense against xenobiotics.