Identifying genes required for digestive physiology and lipid metabolism

肠道对膳食营养素的吸收是导致人类多种代谢性疾病的重要过程。动物对高脂肪食物的反应需要消化组织和肠道微生物区系之间的协调。肠细胞(EC)是肠道的吸收细胞，在含有载脂蛋白B(ApoB)的脂蛋白中为远端组织循环准备脂类。感觉内分泌细胞(EEC)通过钙依赖的激素释放将营养信息传递给其他细胞和组织。然而，介导EC和EEC对高脂肪膳食的餐后反应的转录和信号通路尚不清楚，微生物区系如何影响这些相互作用也是未知的。为了解决这些知识差距，我们的研究团队开创了斑马鱼系统，用于研究脂代谢和宿主-微生物区系相互作用。这包括(1)成像消化器官脂质摄取、运输和储存的新方法，(2)从消失的少量物质中量化含有ApoB的脂蛋白的大小和数量的第一个报告线，(3)允许分析体内对饮食营养的实时反应的EEC活性的荧光指示器，以及(4)经济高效的灵芝操作方法。利用这些工具进行仅在斑马鱼幼体中可能的高分辨率体内成像，我们揭示了EC、EEC和微生物餐后对饮食脂肪的反应背后的动态整合途径。这包括在EC中宿主转录因子和脂蛋白合成酶途径之间的早期餐后相互作用，以及由微生物区系介导的EEC餐后晚期适应性反应。这项建议的目的是确定这些EC和EEC餐后对膳食脂肪的反应的分子机制，以及受微生物区系控制的具体步骤。我们将检验这一中心假设，即微生物区系促进膳食脂肪的脂肪分解为脂肪酸，这些脂肪酸被EC吸收，导致转录程序的激活和含有ApoB的脂蛋白颗粒的合成，这些颗粒被EEC集体感知以改变其活性。这一竞争性续签利用了三个领域领先实验室之间的长期合作伙伴关系，拥有一套强大的突变和新型转基因报告系，这些报告系在上一次资助期间开发。这项拟议研究的预期结果预计将产生重大影响，因为它们可能导致合理操纵EC、EEC和微生物区系的相互作用和对饮食脂肪的反应的新策略，这些策略可用于降低人类代谢性疾病的发生率和严重性。

Intestinal absorption of dietary nutrients is an important process contributing to the etiology of multiple metabolic diseases in humans. An animal’s response to a high-fat meal requires coordination between digestive tissues and intestinal microbiota. Enterocytes (EC) are the absorptive cells of the intestine that prepare lipids for circulation to distal tissues in Apolipoprotein B (ApoB)-containing lipoproteins. Sensory enteroendocrine cells (EEC) communicate nutrient information to other cells and tissues via calcium- dependent hormone release. However, the transcriptional and signaling pathways mediating EC and EEC postprandial responses to a high-fat meal are unclear, and how microbiota influence these interactions is unknown. To address these knowledge gaps, our research team has pioneered the zebrafish system for studies of lipid metabolism and host-microbiota interaction. This includes (1) novel methods to image digestive organ lipid uptake, transport and storage, (2) the first reporter line to quantify the size and numbers of ApoB- containing lipoproteins from vanishing small amounts of material, (3) a fluorescent indicator of EEC activity that permits analysis of in vivo real-time responses to dietary nutrients, and (4) methods for cost-efficient gnotobiotic manipulation. Leveraging these tools for high-resolution in vivo imaging only possible in the larval zebrafish, we have uncovered a dynamic integrative pathway underlying EC, EEC, and microbial postprandial responses to dietary lipids. This includes early postprandial interactions in EC between a host transcription factor and the lipoprotein synthesis enzyme pathway, and late postprandial adaptive responses by EEC that are mediated by microbiota. The objective of this proposal is to define the molecular mechanisms underlying these EC and EEC postprandial responses to dietary lipid, and the specific steps controlled by microbiota. We will test the central hypothesis that microbiota promote lipolysis of dietary fat into fatty acids that are absorbed by EC leading to activation of a transcriptional program and synthesis of ApoB-containing lipoprotein particles, which are collectively perceived by EEC to alter their activity. This competitive renewal leverages long-standing partnerships between three field-leading labs with a powerful set of mutant and novel transgenic reporter lines developed during the prior funding period. The expected outcomes of the proposed research are expected to have a significant impact because they are likely to lead to new strategies for rationally manipulating EC, EEC, and microbiota interactions and responses to dietary fat which could be used to reduce incidence and severity of metabolic diseases in humans.