Genome-wide high resolution map of transcription start sites across the life cycle of Oikopleura dioica. Oikopleura dioica

The selection and activity of transcription start sites (TSSs) are central to gene regulation and depend on sequence and chromatin features within a core promoter region. We present, at single-bp resolution, the TSS landscape of Oikopleura dioica, a marine chordate in the sister group to vertebrates and relate dynamics of promoter usage, and their architectures, across its life cycle, to chromatin features. TSS selection grammars include a genome-wide shift during spermatogenesis to promoters with a novel, position-specific, core promoter element, present in >70% of male-specific promoters. Differential promoter usage in spermatogenesis includes the activation of cryptic internal promoters within polycistronic operons, alternative TSS selection within promoters of genes expressed in earlier development and the exclusion of trans-splice sites. In contrast, maternal promoters, which are active in endocycling nurse nuclei in O. dioica, lack position-specific motifs, have dispersed TSS usage, ordered nucleosomes around the TSS and typical histone modifications associated with broad promoters. Furthermore, our results indicate that the cell cycle regulator, E2F1, acts as a master regulator of these maternally expressed genes. Zygotic promoters are also predominantly broad and the vertebrate maternal to zygotic switch in mode of TSS-selection is absent. DNA methylation in gene bodies is an ancient property of eukaryotic genomes and is thought to repress alternative TSSs. We report a strong association between gene body DNA methylation, H3K4me3-depletion and TATA-dependent sharp promoters in O. dioica. Core promoter features may therefore determine the subset of gene bodies that are methylated in this invertebrate genome. Overall design: RNA was extracted from 6 key stages of development, depleted of trans-spliced transcripts and TSSs profiled using DeepCAGE.

转录起始位点（transcription start sites, TSSs）的选择与活性是基因调控的核心环节，其依赖于核心启动子区域内的序列特征与染色质特征。本研究以单碱基分辨率绘制了住囊虫（Oikopleura dioica）——一种隶属于脊椎动物姊妹群的海洋脊索动物——的转录起始位点全景图谱，并将其整个生命周期内的启动子使用动态及其结构与染色质特征进行关联分析。转录起始位点选择规则显示，在精子发生过程中，全基因组范围内的启动子选择发生转向，偏好携带新型位置特异性核心启动子元件的启动子，这类元件存在于70%以上的雄性特异性启动子中。精子发生过程中的启动子使用差异包括：多顺反子操纵子内隐秘内部启动子的激活、早期发育表达基因启动子内的可变转录起始位点选择，以及反式剪接位点的排除。与之相反，在住囊虫内循环滋养细胞核中活跃的母源性启动子，不具备位置特异性基序，呈现分散式转录起始位点使用模式，其转录起始位点周围排布着规整的核小体，并带有广谱启动子典型的组蛋白修饰特征。此外，本研究结果表明，细胞周期调控因子E2F1是这些母源性表达基因的核心调控因子。合子启动子同样以广谱型为主，且不存在脊椎动物中所见的转录起始位点选择模式的母源-合子转换现象。基因体DNA甲基化是真核生物基因组的古老特征，此前被认为可抑制可变转录起始位点的使用。本研究发现，在住囊虫中，基因体DNA甲基化、H3K4me3缺失与依赖TATA的尖锐型启动子之间存在显著关联。因此，核心启动子特征或许决定了该无脊椎动物基因组中发生甲基化的基因体子集。实验设计：从6个关键发育阶段提取总RNA，去除反式剪接转录本，并利用DeepCAGE技术对转录起始位点进行测序分析。