Department of Cell and Molecular Physiology, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States of America.
PLoS Genet. 2012;8(3):e1002585. doi: 10.1371/journal.pgen.1002585. Epub 2012 Mar 29.
The intestinal microbiota enhances dietary energy harvest leading to increased fat storage in adipose tissues. This effect is caused in part by the microbial suppression of intestinal epithelial expression of a circulating inhibitor of lipoprotein lipase called Angiopoietin-like 4 (Angptl4/Fiaf). To define the cis-regulatory mechanisms underlying intestine-specific and microbial control of Angptl4 transcription, we utilized the zebrafish system in which host regulatory DNA can be rapidly analyzed in a live, transparent, and gnotobiotic vertebrate. We found that zebrafish angptl4 is transcribed in multiple tissues including the liver, pancreatic islet, and intestinal epithelium, which is similar to its mammalian homologs. Zebrafish angptl4 is also specifically suppressed in the intestinal epithelium upon colonization with a microbiota. In vivo transgenic reporter assays identified discrete tissue-specific regulatory modules within angptl4 intron 3 sufficient to drive expression in the liver, pancreatic islet β-cells, or intestinal enterocytes. Comparative sequence analyses and heterologous functional assays of angptl4 intron 3 sequences from 12 teleost fish species revealed differential evolution of the islet and intestinal regulatory modules. High-resolution functional mapping and site-directed mutagenesis defined the minimal set of regulatory sequences required for intestinal activity. Strikingly, the microbiota suppressed the transcriptional activity of the intestine-specific regulatory module similar to the endogenous angptl4 gene. These results suggest that the microbiota might regulate host intestinal Angptl4 protein expression and peripheral fat storage by suppressing the activity of an intestine-specific transcriptional enhancer. This study provides a useful paradigm for understanding how microbial signals interact with tissue-specific regulatory networks to control the activity and evolution of host gene transcription.
肠道微生物群增强了膳食能量的获取，导致脂肪在脂肪组织中积累增加。这种作用部分是由微生物抑制肠上皮细胞表达一种称为血管生成素样 4（Angptl4/Fiaf）的脂蛋白脂肪酶循环抑制剂引起的。为了确定肠道特异性和微生物控制 Angptl4 转录的顺式调节机制，我们利用了斑马鱼系统，其中宿主调节 DNA 可以在活体、透明和无菌的脊椎动物中快速分析。我们发现，斑马鱼 angptl4 在包括肝脏、胰岛和肠上皮在内的多种组织中转录，这与其哺乳动物同源物相似。斑马鱼 angptl4 也在定植微生物群后特异性地在肠上皮细胞中被抑制。体内转基因报告基因测定鉴定了 angptl4 内含子 3 内的离散组织特异性调节模块，足以驱动肝脏、胰岛β细胞或肠上皮细胞中的表达。来自 12 种硬骨鱼的 angptl4 内含子 3 序列的比较序列分析和异源功能测定显示，胰岛和肠调节模块的进化存在差异。高分辨率功能图谱和定点突变分析确定了肠活性所需的最小调节序列集。引人注目的是，微生物群抑制了肠特异性调节模块的转录活性，类似于内源性 angptl4 基因。这些结果表明，微生物群可能通过抑制肠特异性转录增强子的活性来调节宿主肠道 Angptl4 蛋白表达和外周脂肪储存。这项研究为理解微生物信号如何与组织特异性调节网络相互作用以控制宿主基因转录的活性和进化提供了一个有用的范例。
