Abstract：
Eukaryotic chromosomes are divided into domains with distinct structural and functional properties, such as differing levels of chromatin compaction and gene transcription. Domains of relatively compact chromatin and minimal transcription are termed heterochromatic, whereas euchromatin is more open and actively transcribed. Insulators separate these domains and maintain their distinct features. Disruption of insulators can cause diseases such as cancer. Many insulators contain tRNA genes (tDNAs), examples of which have been shown to block the spread of activating or silencing activities. This characteristic of specific tDNAs is conserved through evolution, such that human tDNAs can serve as barriers to the spread of silencing in fission yeast. Here we demonstrate that tDNAs from the methylotrophic fungus Pichia pastoris can function effectively as insulators in distantly-related budding yeast. Key to the function of tDNAs as insulators is TFIIIC, a transcription factor that is also required for their expression. TFIIIC binds additional loci besides tDNAs, some of which have insulator activity. Although the mechanistic basis of TFIIIC-based insulation has been studied extensively in yeast, it is largely uncharacterized in metazoa. Utilising publicly-available genome-wide ChIP-seq data, we consider the extent to which mechanisms conserved from yeast to man may suffice to allow efficient insulation by TFIIIC in the more challenging chromatin environments of metazoa and suggest features that may have been acquired during evolution to cope with new challenges. We demonstrate the widespread presence at human tDNAs of USF1, a transcription factor with well-established barrier activity in vertebrates. We predict that tDNA-based insulators in higher organisms have evolved through incorporation of modules, such as binding sites for factors like USF1 and CTCF that are absent from yeasts, thereby strengthening function and providing opportunities for regulation between cell types.
摘要：
真核染色体分为具有不同结构和功能特性的结构域，例如不同水平的染色质压缩和基因转录。相对紧凑的染色质和最小转录的域被称为异色，而常染色质更开放且转录活跃。绝缘体将这些域分开并保持其独特的特征。绝缘体的破坏会导致癌症等疾病。许多绝缘子含有tRNA基因（tDNA），其中的例子已被证明可以阻止激活或沉默活动的传播。特定tDNA的这种特征通过进化是保守的，这样人类tDNA可以作为裂变酵母沉默传播的屏障。在这里，我们证明了来自甲基营养真菌巴斯德毕赤酵母的tDNA可以有效地充当远缘相关的出芽酵母的绝缘体。tDNA作为绝缘体功能的关键是TFIIIC，转录因子，也是其表达所必需的。TFIIIC结合除tDNA之外的其他基因座，其中一些有绝缘体活动。尽管已经在酵母中广泛研究了基于TFIIIC的绝缘的机理基础，它在后生中基本上没有特征。利用公开可用的全基因组ChIP-seq数据，我们考虑了从酵母到人的保守机制可能足以在后生动物更具挑战性的染色质环境中通过TFIIIC进行有效隔离的程度，并提出了在进化过程中可能获得的特征以应对新的挑战。我们证明了USF1在人类tDNA中的广泛存在，USF1是一种在脊椎动物中具有良好屏障活性的转录因子。我们预测，高等生物中基于tDNA的绝缘体通过模块的掺入而进化，例如酵母中不存在的USF1和CTCF等因子的结合位点，从而加强功能并为细胞类型之间的调节提供机会。
