Drosophila non-gonadal PIWI protein Aubergine regulates regenerative stem cell proliferation and tumourigenesis in the adult intestine [RNA-seq]

Somatic stem cells are executors of physiological and pathological proliferation of adult self-renewing tissues, such as the intestine. The actions of intestinal stem cells (ISCs) rely on the integration of cell intrinsic and niche-derived signals, which are necessary to achieve a balanced response to the multiple stimuli that constantly challenge tissue homeostasis and organismal health. Disruption of such balance is causative of age-associated tissue dysfunction and hyperproliferative conditions, including inflammation and cancer. The highly conserved PIWI-interacting RNAs (piRNAs) biosynthesis pathway, also known as the PIWI pathway, has been classically studied in the Drosophila germline for its role in the repression of transposable elements (TEs) and the regulation of germline stem cell homeostasis. Recent reports have emerged on a role of Piwi, the founding member of the pathway, in the maintenance of ISC homeostasis in the adult Drosophila midgut. The implications of these findings regarding a general role of the PIWI pathway and piRNAs in the intestine remain to be addressed. Here, we characterise a cell autonomous role of the PIWI family protein Aubergine (Aub) in ISCs. We show that, while dispensable for homeostatic self-renewal of the midgut, inducible Aub is essential to regulate ISC proliferation following acute damage of the intestinal epithelium and in oncogenic settings. Our work suggests that the role of Aub in ISCs is independent of its piRNAs regulatory function. Instead, Aub drives ISC proliferation through the induction of regenerative stem cell factors Myc and Sox21a, and through the initiation of protein translation by the eukaryotic initiation factor 3 complex (eIF3). In summary, our results discover a role of Aub in damage induced proliferation of the adult Drosophila intestine involving the regulation of major regenerative signalling pathways and the protein translation machinery. Furthermore, we present genetically distinct hyperplastic settings fostering a role of Aub as an intestinal tumour promoter or suppressor. The study aims at understanding the role of the PIWI family protein Aubergine in the proliferation of intestinal stem cells (ISCs) upon damage in Drosophila melanogaster. We generated oxidised small RNA sequencing libraries from freshly dissected midguts, sorted ISCs/Enteroblasts (marked by escargot-Gal4) and ovaries. We also generated RNA sequencing libraries from poly-A plus RNA to determine the abundance of transposon mRNAs expressed in the ISCs/Enteroblasts.

体细胞干细胞（somatic stem cells）是成年自我更新组织（如肠道）生理与病理性增殖的核心执行者。肠道干细胞（intestinal stem cells, ISCs）的功能依赖于细胞内在信号与微环境来源信号的整合，这是对持续挑战组织稳态与机体健康的多种刺激做出平衡应答的必要条件。这种平衡的破坏会引发与衰老相关的组织功能障碍及增殖异常状态，包括炎症与癌症。高度保守的PIWI互作RNA（PIWI-interacting RNAs, piRNAs）生物合成通路，又称PIWI通路，经典上以果蝇生殖系为研究模型，其功能为转座因子（transposable elements, TEs）沉默以及生殖系干细胞稳态调控。已有最新研究表明，该通路的创始成员Piwi可参与成年果蝇中肠的肠道干细胞稳态维持。上述发现关于PIWI通路与piRNAs在肠道中的通用功能的相关启示，仍有待进一步阐释。本研究中，我们解析了PIWI家族蛋白Aubergine（Aub）在肠道干细胞中的细胞自主性功能。我们的研究表明，尽管Aub对于中肠的稳态自我更新并非必需，但诱导性表达的Aub在肠道上皮急性损伤后以及致癌环境中，对肠道干细胞增殖的调控至关重要。我们的研究结果提示，Aub在肠道干细胞中的功能与其调控piRNAs的功能相互独立。相反，Aub通过诱导再生干细胞因子Myc与Sox21a的表达，并借助真核翻译起始因子3复合物（eukaryotic initiation factor 3 complex, eIF3）启动蛋白质翻译过程，从而促进肠道干细胞的增殖。综上，本研究结果揭示了Aub在果蝇成年肠道损伤诱导增殖中的功能，该功能涉及对核心再生信号通路与蛋白质翻译机器的调控。此外，我们构建了遗传背景各异的增生异常模型，证实Aub可作为肠道肿瘤的促进因子或抑制因子发挥作用。本研究旨在探究黑腹果蝇（Drosophila melanogaster）中PIWI家族蛋白Aubergine在损伤诱导的肠道干细胞增殖过程中的功能。我们从新鲜解剖的中肠、分选得到的肠道干细胞/肠母细胞（以escargot-Gal4标记）以及卵巢中构建了氧化型小RNA测序文库。我们同时从poly-A富集RNA中构建了RNA测序文库，以定量检测肠道干细胞/肠母细胞中表达的转座子mRNA丰度。