Identifying genes required for digestive physiology and lipid metabolism

膳食营养物质的肠道吸收是人类多种代谢性疾病病因的重要过程。动物对高脂肪食物的反应需要消化组织和肠道微生物群之间的协调。肠细胞（EC）是肠道的吸收细胞，它以载脂蛋白B （ApoB）为载体，为远端组织的循环准备脂质。感觉肠内分泌细胞（EEC）通过钙依赖性激素的释放将营养信息传递给其他细胞和组织。然而，介导EC和EEC对高脂肪膳食餐后反应的转录和信号通路尚不清楚，微生物群如何影响这些相互作用也尚不清楚。为了解决这些知识空白，我们的研究团队率先在斑马鱼系统中研究脂质代谢和宿主-微生物群相互作用。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ApoB脂蛋白的大小和数量的第一个报告线，(3)EEC活性的荧光指示器，允许分析体内对膳食营养素的实时反应，以及(4)具有成本效益的生物操作方法。利用这些工具对斑马鱼幼虫进行高分辨率体内成像，我们发现了EC、EEC和微生物对膳食脂质的餐后反应背后的动态综合途径。这包括早期餐后EC中宿主转录因子与脂蛋白合成酶途径之间的相互作用，以及由微生物群介导的晚餐后EC的适应性反应。本提案的目的是确定这些EC和EEC对膳食脂质的餐后反应的分子机制，以及微生物群控制的具体步骤。我们将验证微生物群促进膳食脂肪分解为脂肪酸的中心假设，这些脂肪酸被EC吸收，导致转录程序的激活和含载脂蛋白的脂蛋白颗粒的合成，这些颗粒被EEC共同感知以改变其活性。这一竞争性更新利用了三个领域领先的实验室之间的长期合作伙伴关系，这些实验室在先前的资助期间开发了一套强大的突变和新型转基因报告系。这项拟议研究的预期结果预计将产生重大影响，因为它们可能会导致合理控制EC、EEC和微生物群相互作用以及对膳食脂肪反应的新策略，从而可用于降低人类代谢疾病的发病率和严重程度。

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.