Molecular basis of host-microbiota signaling in the zebrafish gut

描述(申请人提供)：所有动物都在微生物丰富的环境中进化，其中许多微生物以亲密的组合形式存在，塑造了动物宿主的发育和正常生理。我们建立了一个无菌(GF)斑马鱼模型来研究本土微生物群在肠道发育中的作用。我们发现，斑马鱼的消化道表现出多种发育缺陷，包括不表达成熟的碱性磷酸酶(IAP)活性和含有Gala1，3-半乳糖的多糖。我们已经证明，纯化的脂多糖(LPS)足以在GF动物体内诱导IAP活性。相反，对半乳糖结合体的调节需要用本土微生物区系的成分来定居。我们进一步证明，这两条独立的途径，使用不同的微生物信号，通过IAP和半乳糖基转移酶(GATIT)基因的转录调节来调节宿主的反应。 在这项资助中，我们将研究寄主和其微生物区系之间这两条信号通路的分子基础和功能意义。我们的初步数据表明，IAP的活性被证明是去磷酸化的内毒素，在肠道中是需要的，以防止内毒素的毒性。我们将测试这种肠道酶的正常功能是对革兰氏阴性共生菌和致病菌的解毒和减少肠道炎症的假设。为了研究斑马鱼半乳糖偶联体表达的功能和微生物调控，我们建立了斑马鱼与一种遗传易驯化细菌Veronas biovar sobria的单关联模型。我们将使用细菌遗传和斑马鱼转基因方法来识别调节GalT转录的微生物信号和宿主途径。最后，我们将检验这一假设，即肠道Gala1，3Gal多糖的表达促进了适当微生物区系的建立。 拟议的研究将教给我们关于我们与微生物区系接触的分子对话，并将阐明这种对话在Gl炎症性疾病、机会性感染和败血症等疾病中被破坏的方式。

DESCRIPTION (provided by applicant): All animals have evolved in an environment rich in microorganisms, many of which exist in intimate associations that shape the animal hosts' development and normal physiology. We have established a germfree (GF) zebrafish model to study the role of the indigenous microbiota in gut development. We find that the GF zebrafish digestive tract exhibits multiple developmental defects, including failure to express mature patterns of alkaline phosphatase (IAP) activity and Gala1,3Gal containing glycans. We have demonstrated that purified lipopolysaccharide (LPS) is sufficient to induce IAP activity in GF animals. In contrast, regulation of the galactoconjugate requires colonization with constituents of the indigenous microbiota. We have further shown that these two independent pathways, using different microbial signals, regulate host responses through the transcriptional regulation of the IAP and galactosyltransferase (GaIT) genes. In this grant we will investigate the molecular basis and functional significance of these two signaling pathways between the host and its microbiota. Our preliminary data demonstrate that IAP activity, which has been shown to dephosphorylated LPS, is required in the intestine to protect against LPS toxicity. We will test the hypothesis that the normal function of this intestinal enzyme is to detoxifies LPS and reduces intestinal inflammation in response to gram-negative commensal and pathogenic bacteria. To investigate the function and microbial regulation of galactoconjugate expression in the zebrafish we have established a zebrafish mono-association model with a genetically tractable bacterium, Aeromonas veronii biovar sobria. We will use bacterial genetic and zebrafish transgenic approaches to identify the microbial signals and the host pathways that regulate GalT transcription. Finally, we will test the hypothesis that intestinal Gala1,3Gal glycan expression promotes the establishment of the appropriate microbiota. The proposed research will teach us about the molecular dialogues in which we engage with our microbiota and will shed light on the ways in which this dialogue is disrupted in diseases such as Gl inflammatory disorders, opportunistic infections, and sepsis.