Chitin synthesizing supercomplexes (ChiSC) in insects: analysis of constituents and functions

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Chitin nanofibers are highly versatile structural components of different types of extracellular matrices including insect cuticles and peritrophic matrices. Proteins involved in the biosynthesis of chitin are attractive targets for the development of eco-friendly insecticides. However, the development of this research field is still in its early stage due to the lack of knowledge on the precise composition of chitin-synthesizing complexes and the function of proteins necessary for chitin formation particularly at post-catalytic steps. We hypothesize that chitin biogenesis depends on chitin synthesizing supercomplexes (ChiSC), which combine different non-catalytic and catalytic functions that are required to orchestrate chitin nanofibril formation. Our previous work suggests that specific chitinases, chitin-binding proteins, chitin deacetylases (involved in chitosan formation), and possibly ABC transporters physically interact with the chitin synthase during chitin biogenesis. We have already started to explore the structure, function and regulation of some of these candidates of ChiSC constituents. To learn about the core components of the insect ChiSC, we will perform a comparative study using two phylogenetically distant but well-established model insect pests for chitin biosynthesis, Tribolium castaneum and Locusta migratoria. In an joint research effort of the Chinese and German partners, the following four objectives are addressed: 1) Identification of novel ChiSC constituents and analyses of ChiSC localization, assembly and disassembly; 2) Characterization of the function of each ChiSC component and its influence on chitin nanofibril organization in elytral cuticles; 3) Functional co-expression of CHS and ChiSC constituents, and biochemical analysis of CHS activity and interfacial interactions; 4) High-speed AFM analyses of the effects of individual ChiSC constituents and mutagenized versions on the kinetics of chitin nanofibril formation. The expected outcomes will not only add new knowledge to the composition of the basal machinery of chitin biosynthesis and its mode of action in insects, but may also reveal potential targets for developing novel insecticides interfering with chitin synthesis.