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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Functional cis-regulatory genomics for systems biology
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Functional cis-regulatory genomics for systems biology

机译：系统生物学的功能顺式调控基因组学

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Gene expression is controlled by interactions between trans-regulatory factors and cis-regulatory DNA sequences, and these interactions constitute the essential functional linkages of gene regulatory networks (GRNs). Validation of GRN models requires experimental as-regulatory tests of predicted linkages to authenticate their identities and proposed functions. However, cis-regulatory analysis is, at present, at a severe bottleneck in genomic system biology because of the demanding experimental methodologies currently in use for discovering cis-regulatory modules (CRMs), in the genome, and for measuring their activities. Here we demonstrate a high-throughput approach to both discovery and quantitative characterization of CRMs. The unique aspect is use of DNA sequence tags to "barcode" CRM expression constructs, which can then be mixed, injected together into sea urchin eggs, and subsequently deconvolved. This method has increased the rate of as-regulatory analysis by > 100-fold compared with conventional one-by-one reporter assays. The utility of the DNA-tag reporters was demonstrated by the rapid discovery of 81 active CRMs from 37 previously unexplored sea urchin genes. We then obtained simultaneous high-resolution temporal characterization of the regulatory activities of more than 80 CRMs. On average 2-3 CRMs were discovered per gene. Comparison of endogenous gene expression profiles with those of the CRMs recovered from each gene showed that, for most cases, at least one CRM is active in each phase of endogenous expression, suggesting that CRM recovery was comprehensive. This approach will qualitatively alter the practice of GRN construction as well as validation, and will impact many additional areas of regulatory system biology.
机译：基因表达受反式调节因子和顺式调节性DNA序列之间相互作用的控制，这些相互作用构成了基因调节网络（GRN）的基本功能连接。要验证GRN模型，需要对预测的链接进行实验性的调节性测试，以验证其身份和提出的功能。然而，由于目前用于发现基因组中的顺式调控模块（CRM）以及测量其活性的严格的实验方法，顺式调控分析目前是基因组系统生物学中的严重瓶颈。在这里，我们展示了一种用于CRM的发现和定量表征的高通量方法。独特的方面是使用DNA序列标签“编码” CRM表达构建体，然后可以将其混合，一起注入海胆卵中，然后进行去卷积。与常规的一对一报告基因检测方法相比，此方法将调节性分析的速度提高了100倍以上。通过从37个以前未开发的海胆基因中快速发现81个活性CRM，证明了DNA标签报道基因的实用性。然后，我们获得了80多个CRM的监管活动的同步高分辨率时间特征。每个基因平均发现2-3个CRM。内源基因表达谱与从每个基因回收的CRM的表达谱的比较表明，在大多数情况下，内源表达的每个阶段至少有一个CRM在活性，这表明CRM的回收是全面的。这种方法将在质量上改变GRN构建和验证的实践，并将影响监管体系生物学的许多其他领域。

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