CONSTITUTIVE EXPRESSION OF DIGESTIVE CYSTEINE PROTEINASE FORMS DURING DEVELOPMENT OF THE COLORADO POTATO BEETLE, LEPTINOTARSA-DECEMLINEATA SAY (COLEOPTERA, CHRYSOMELIDAE)

Changes in digestive proteinase activities were monitored throughout development of the Colorado potato beetle (CPB), Leptinotarsa decemlineata Say. Consistent with high growth rates generally observed for larval stages, larva midgut extracts exhibited high specific proteinase activity as compared with adult extracts. Apart from these quantitative variations, no qualitative difference was noted between developmental stages. Specific activity was maximal at mildly acidic pH, and was activated by reducing agents and phenylmethylsulfonyl fluoride. The activity was significantly decreased by trans-epoxysuccinyl-L-leucylamido (4-guanidino) butane and pepstatin, indicating the presence of cysteine and aspartate proteinase activity in the extracts, respectively. As demonstrated by the use of class-specific activity gels, this activity was principally due to the constitutive expression of at least 9 cysteine proteinase forms active over a wide pH range. In accordance with the apparent existence of a unique digestive proteolytic system throughout CPB development, the rice cysteine proteinase inhibitor oryzacystatin I had the same inhibitory effect on extracts of each developmental stage, but in contrast to early stages, growth of 3rd and 4th instars was not significantly affected when the inhibitor was fed to the insects. These observations indicate the potential of the rice cystatin for inhibition of CPB growth but also suggest a stage-related efficiency for this control strategy, independent of the nature of proteinase forms used by the insect.