Most insects possess two different acetylcholinesterases (AChE1 and AChE2) but it remains unknown which AChE plays the major role in synaptic transmission. To investigate the evolutionary distribution of AChE1 and AChE2, the AChE with the main catalytic function in several insect species belonging to 18 representative orders was determined by native-PAGE in conjunction with Western blotting using AChE1- and AChE2-specific antibodies. Among the 98 insect species examined, AChE1 was expressed as the main enzyme in 65 species across diverse taxa. In the remaining species, however, AChE2 was expressed as the major enzyme. These findings are contrary to the common expectation that AChE1 is major enzyme in most insects, with the exception of Cyclorrhapha, and further demonstrate that the specialization of AChE2 as the main enzyme or the replacement of AChE1 function with AChE2 function were rather common events, having multiple independent origins during insect evolution. It was hypothesized that the generation of multiple AChE2 isoforms via alternative splicing has allowed the loss of ace1during the functional replacement of AChE1 with AChE2 in Cyclorrhapha. However, the presence of AChE2 as the main AChE in higher social Hymenoptera provides an example of the functional replacement of AChE1 with AChE2 without the loss of ace1. The current findings should provide valuable insights into which AChE has evolved to perform synaptic functions and to become the main insecticide target in different species

• Young Ho Kim(Research Institute for Agriculture and Life Sciences, Seoul National University)
• Si Hyeock Lee(Research Institute for Agriculture and Life Sciences, Seoul National University, Department of Agricultural Biotechnology, Seoul National University)