High-throughput cis-regulatory element discovery in the vector mosquito Aedes aegypti
High-throughput cis-regulatory element discovery in the vector mosquito Aedes aegypti
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DOI：
10.1186/s12864-016-2468-x
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发表时间：
2016-05-10
期刊：
影响因子：
4.4
通讯作者：
Duman-Scheel, Molly
中科院分区：
文献类型：
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作者：
Behura, Susanta K.;Sarro, Joseph;Duman-Scheel, Molly
背景：尽管蚊子基因组和遗传学研究取得了实质性进展，但在蚊子或其他非模式昆虫中几乎没有发现顺式调控元件(Cres)，即控制基因表达的DNA序列。甲醛辅助分离调控元件，结合DNA测序，FIRE-SEQ，正在成为发现全球CRE的一种强大的新的高通量工具。FIRE导致不受核小体结合的开放染色质DNA片段的优先回收，核小体是一种进化上保守的调节活性指标，然后对其进行测序。尽管这种方法很强大，但FIRE-SEQ尚未被应用于非模型昆虫的研究。在这项研究中，我们利用FIRE-SEQ来分析开放染色质，并确定人类疾病媒介埃及伊蚊基因组中可能的调控元件。结果：通过对从埃及伊蚊胚胎中分离的FIRE DNA的下一代测序获得的序列数据分析，发现了121,000个FIRE峰(FP)，其中许多聚集在已知在该阶段表达的基因的1kb的5‘上游侧翼区。正如预期的那样，已知的转录因子共识结合位点在FFP中得到了丰富，其中FoxA1、驼背、GFI、KLF4、MYB/PH3和Sox9最占优势。在转基因果蝇的报告分析中，所有在体内测试的元素都被证实可以驱动基因表达。在最近在登革热病毒敏感和耐药的蚊子株中发现的13,000个单核苷酸多态(SNPs)中，3365个被发现映射到FP。结论：通过对埃及伊蚊开放染色质的FIRE-SEQ分析，可以在全基因组范围内发现Cres。这项研究的结果表明，FIRE-SEQ是鉴定非模式生物基因组中调控DNA的有效工具，包括人类疾病媒介蚊子。
Background: Despite substantial progress in mosquito genomic and genetic research, few cis-regulatory elements (CREs), DNA sequences that control gene expression, have been identified in mosquitoes or other non-model insects. Formaldehyde-assisted isolation of regulatory elements paired with DNA sequencing, FAIRE-seq, is emerging as a powerful new high-throughput tool for global CRE discovery. FAIRE results in the preferential recovery of open chromatin DNA fragments that are not bound by nucleosomes, an evolutionarily conserved indicator of regulatory activity, which are then sequenced. Despite the power of the approach, FAIRE-seq has not yet been applied to the study of non-model insects. In this investigation, we utilized FAIRE-seq to profile open chromatin and identify likely regulatory elements throughout the genome of the human disease vector mosquito Aedes aegypti. We then assessed genetic variation in the regulatory elements of dengue virus susceptible (Moyo-S) and refractory (Moyo-R) mosquito strains.Results: Analysis of sequence data obtained through next generation sequencing of FAIRE DNA isolated from A. aegypti embryos revealed >121,000 FAIRE peaks (FPs), many of which clustered in the 1 kb 5' upstream flanking regions of genes known to be expressed at this stage. As expected, known transcription factor consensus binding sites were enriched in the FPs, and of these FoxA1, Hunchback, Gfi, Klf4, MYB/ph3 and Sox9 are most predominant. All of the elements tested in vivo were confirmed to drive gene expression in transgenic Drosophila reporter assays. Of the >13,000 single nucleotide polymorphisms (SNPs) recently identified in dengue virus-susceptible and refractory mosquito strains, 3365 were found to map to FPs.Conclusion: FAIRE-seq analysis of open chromatin in A. aegypti permitted genome-wide discovery of CREs. The results of this investigation indicate that FAIRE-seq is a powerful tool for identification of regulatory DNA in the genomes of non-model organisms, including human disease vector mosquitoes.