<i>De novo</i> transcriptome analysis of the excretory tubules of <i>Carausius morosus</i> (Phasmatodea) and possible functions of the midgut ‘appendices’

The Malpighian tubules are the insect excretory organs, responsible for ion and water homeostasis and elimination of nitrogenous wastes. Post-genomic assays suggest they also metabolize and detoxify xenobiotic compounds and have antimicrobial properties. The Phasmatodea have an additional, unique set of excretory organs referred to predominantly as midgut appendices. Their function and how it compares to phasmid and other insect Malpighian tubules is unknown. Hypotheses include carbonic anhydrase activity, calcium and metal cation sequestration, and xenobiotic transport. This work presents the first comparative transcriptomic analysis of the Phasmatodean excretory organs, using the model insect Carausius morosus. I produced de novo transcriptomes of the midgut appendices, midgut wall, and Malpighian tubules, and looked for differentially expressed genes associated with putative organ functions. The appendices differentially and highly express lipid transport and metabolism proteins, and the biomineralization gene otopetrin. The Malpighian tubules differentially and highly express acid phosphatases and multiple transporter types, while appendices express fat-soluble vitamin and peptide transporters. Many defense proteins such as multidrug resistance proteins, ABC transporters, cytochrome P450’s, and glutathione-S-transferases were differentially expressed in specific excretory organs. I hypothesize that the appendices and Malpighian tubules both have defensive / xenobiotic metabolism functions, but each likely target different substrates. Phasmid Malpighian tubules excrete as in other insects, while the appendices may predominantly regulate amino acids, fats, and fat-soluble compounds. Lipid metabolism in insects is poorly understood, and the Phasmatodea may thus serve as a model for studying this further.马氏管（Malpighian tubules）是昆虫的排泄器官，负责维持离子与水稳态，并清除含氮废物。后基因组学分析表明，这类器官还可代谢并解毒外源性化合物，同时具备抗菌活性。竹节虫目（Phasmatodea）拥有一套额外的独特排泄器官，目前主要被称为中肠附属物（midgut appendices）。目前对于该附属物的功能，以及其与竹节虫及其他昆虫马氏管的功能差异尚不明确。现有假说认为其可能具备碳酸酐酶活性、钙与金属离子螯合能力，以及外源性化合物转运功能。 本研究以模式昆虫Carausius morosus为材料，首次对竹节虫目昆虫的排泄器官开展比较转录组学分析。我们构建了中肠附属物、中肠壁以及马氏管的从头转录组（de novo transcriptomes），并筛选了与各器官推定功能相关的差异表达基因（differentially expressed genes）。结果显示，中肠附属物显著高表达脂质转运与代谢蛋白，以及生物矿化基因otopetrin；马氏管则显著高表达酸性磷酸酶与多种转运蛋白类型，而中肠附属物则高表达脂溶性维生素与肽转运蛋白。多种防御蛋白，如多药耐药蛋白（multidrug resistance proteins）、ABC转运蛋白（ABC transporters）、细胞色素P450酶系（cytochrome P450s）以及谷胱甘肽S-转移酶（glutathione-S-transferases），在特定排泄器官中呈现差异表达。 我们据此提出假说：中肠附属物与马氏管均具备防御/外源性化合物代谢功能，但二者靶向的底物可能存在差异。竹节虫的马氏管与其他昆虫一样行使排泄功能，而中肠附属物则可能主要负责调控氨基酸、脂肪及脂溶性化合物的稳态。目前对于昆虫脂质代谢的研究尚不完善，因此竹节虫目可作为进一步研究脂质代谢的理想模型。