Partially redundant enhancers cooperatively maintain Mammalian pomc expression above a critical functional threshold.
Partially redundant enhancers cooperatively maintain Mammalian pomc expression above a critical functional threshold.
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DOI：
10.1371/journal.pgen.1004935
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发表时间：
2015-02
期刊：
影响因子：
4.5
通讯作者：
Low MJ
中科院分区：
文献类型：
--
作者：
Lam DD;de Souza FS;Nasif S;Yamashita M;López-Leal R;Otero-Corchon V;Meece K;Sampath H;Mercer AJ;Wardlaw SL;Rubinstein M;Low MJ
许多基因的细胞特异性表达由多个增强子传递，每个增强子控制特定的表达域。相比之下，多个增强子驱动一些参与胚胎发育的基因的相似表达模式，表明调控冗余。果蝇研究表明，功能重叠的增强子通过缓冲基因表达免受环境和遗传干扰来促进发育。然而，人们对脊椎动物和主要在成年期表达的基因的调控冗余知之甚少。在这里，我们研究 nPE1 和 nPE2，这是两种系统发育上保守的哺乳动物增强子，它们驱动阿片黑皮质素原基因 (Pomc) 表达到同一组下丘脑神经元。同时删除两个增强子会消除所有年龄段的 Pomc 表达，并诱导严重的代谢功能障碍，包括早发性极度肥胖。 nPE1 或 nPE2 的靶向失活导致早期胚胎发育过程中 Pomc 表达水平非常低，表明两种增强子具有协同作用。然而，在成年小鼠中，Pomc 表达受到两个增强子的附加控制，其中 nPE1 负责 Pomc 转录的约 80%，nPE2 负责约 20%。因此，nPE1 敲除小鼠表现出轻度肥胖，而 nPE2 缺陷小鼠则保持正常体重。这些结果表明，nPE2 驱动的 Pomc 表达在发育后期被 nPE1 补偿，基本上挽救了 nPE2 缺陷的早期表型。总之，这些结果表明，增强子之间的合作相互作用赋予 Pomc 表达对抗基因调控干扰的稳健性，并排除成年期 Pomc 缺陷引起的有害代谢表型。因此，我们的研究表明，增强子冗余可以被控制哺乳动物成年生理学的基因所利用，并强调了调控序列突变在常见疾病中的潜在意义。面对遗传和环境变化，动物形态和功能的稳定性依赖于一致的基因表达。多个增强子，每个指定一个独特的调节域，控制许多基因的精确时空表达。然而，在一些基因中，明显冗余的增强子以重叠的细胞特异性模式调节表达。尽管这种排列已被证明对无脊椎动物的发育稳健性很重要，但对于脊椎动物物种中明显冗余的增强子以及在成年期发挥作用的基因中的作用却知之甚少。在这里，我们证明哺乳动物 Pomc 基因的表达是由两个这样的明显冗余的增强子以组织特异性方式控制的。我们使用单个增强子的定向删除来描述它们各自对大脑中 Pomc 表达的贡献。由于 Pomc 完整基因座的表达超过了胚胎小鼠中单个增强子对 Pomc mRNA 水平贡献的总和，因此我们推断发育过程中增强子之间存在协同作用。相反，增强子之间的相互作用在成年小鼠中是相加的。同时删除两个增强子几乎完全消除了 Pomc 表达，突变小鼠表现出极度肥胖和代谢功能障碍，而删除单个增强子则具有中等或没有表型效应。总之，我们的结果表明，两个增强子协同维持 Pomc 表达高于关键功能阈值。
Cell-specific expression of many genes is conveyed by multiple enhancers, with each individual enhancer controlling a particular expression domain. In contrast, multiple enhancers drive similar expression patterns of some genes involved in embryonic development, suggesting regulatory redundancy. Work in Drosophila has indicated that functionally overlapping enhancers canalize development by buffering gene expression against environmental and genetic disturbances. However, little is known about regulatory redundancy in vertebrates and in genes mainly expressed during adulthood. Here we study nPE1 and nPE2, two phylogenetically conserved mammalian enhancers that drive expression of the proopiomelanocortin gene (Pomc) to the same set of hypothalamic neurons. The simultaneous deletion of both enhancers abolished Pomc expression at all ages and induced a profound metabolic dysfunction including early-onset extreme obesity. Targeted inactivation of either nPE1 or nPE2 led to very low levels of Pomc expression during early embryonic development indicating that both enhancers function synergistically. In adult mice, however, Pomc expression is controlled additively by both enhancers, with nPE1 being responsible for ∼80% and nPE2 for ∼20% of Pomc transcription. Consequently, nPE1 knockout mice exhibit mild obesity whereas nPE2-deficient mice maintain a normal body weight. These results suggest that nPE2-driven Pomc expression is compensated by nPE1 at later stages of development, essentially rescuing the earlier phenotype of nPE2 deficiency. Together, these results reveal that cooperative interactions between the enhancers confer robustness of Pomc expression against gene regulatory disturbances and preclude deleterious metabolic phenotypes caused by Pomc deficiency in adulthood. Thus, our study demonstrates that enhancer redundancy can be used by genes that control adult physiology in mammals and underlines the potential significance of regulatory sequence mutations in common diseases. The stability of animal form and function in the face of genetic and environmental variation relies on consistent gene expression. Multiple enhancers, each specifying a unique regulatory domain, control the precise spatiotemporal expression of many genes. However, in some genes apparently redundant enhancers regulate expression in overlapping cell-specific patterns. Although this arrangement has been shown to be important for developmental robustness in invertebrates, the role of apparently redundant enhancers in vertebrate species and in genes functioning in adulthood is poorly understood. Here, we show that expression of the mammalian Pomc gene is controlled in a tissue-specific manner by two such apparently redundant enhancers. We used targeted deletion of the individual enhancers to delineate their respective contributions to Pomc expression in the brain. Since Pomc expression from its intact locus exceeds the sum of the individual enhancer contributions to Pomc mRNA levels in embryonic mice, we infer a synergistic action between the enhancers during development. In contrast, the interaction between the enhancers is additive in adult mice. Deletion of both enhancers simultaneously almost completely abolished Pomc expression and the mutant mice displayed extreme obesity and metabolic dysfunction, while deletion of the individual enhancers had a modest or no phenotypic effect. Together, our results demonstrate that the two enhancers cooperatively maintain Pomc expression above a critical functional threshold.
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