8.2% of the Human Genome Is Constrained: Variation in Rates of Turnover across Functional Element Classes in the Human Lineage

Ten years on from the finishing of the human reference genome sequence, it remains unclear what fraction of the human genome confers function, where this sequence resides, and how much is shared with other mammalian species. When addressing these questions, functional sequence has often been equated with pan-mammalian conserved sequence. However, functional elements that are short-lived, including those contributing to species-specific biology, will not leave a footprint of long-lasting negative selection. Here, we address these issues by identifying and characterising sequence that has been constrained with respect to insertions and deletions for pairs of eutherian genomes over a range of divergences. Within noncoding sequence, we find increasing amounts of mutually constrained sequence as species pairs become more closely related, indicating that noncoding constrained sequence turns over rapidly. We estimate that half of present-day noncoding constrained sequence has been gained or lost in approximately the last 130 million years (half-life in units of divergence time, d1/2 = 0.25–0.31). While enriched with ENCODE biochemical annotations, much of the short-lived constrained sequences we identify are not detected by models optimized for wider pan-mammalian conservation. Constrained DNase 1 hypersensitivity sites, promoters and untranslated regions have been more evolutionarily stable than long noncoding RNA loci which have turned over especially rapidly. By contrast, protein coding sequence has been highly stable, with an estimated half-life of over a billion years (d1/2 = 2.1–5.0). From extrapolations we estimate that 8.2% (7.1–9.2%) of the human genome is presently subject to negative selection and thus is likely to be functional, while only 2.2% has maintained constraint in both human and mouse since these species diverged. These results reveal that the evolutionary history of the human genome has been highly dynamic, particularly for its noncoding yet biologically functional fraction.自人类参考基因组序列（human reference genome sequence）完成至今已有十载，人类基因组中具备功能的序列占比、其在基因组中的定位，以及与其他哺乳动物共享的序列比例，仍未明确。在探讨这些问题时，功能性序列常被等同于泛哺乳动物保守序列（pan-mammalian conserved sequence）。然而，那些寿命较短的功能性元件——包括参与物种特异性生物学过程的元件——不会留下长期负选择（negative selection）的印记。本研究通过识别并表征不同分化水平下的真兽类（eutherian）基因组对中受插入缺失约束的序列，解决了上述问题。在非编码序列（noncoding sequence）中，我们发现随着物种对亲缘关系愈发紧密，相互约束序列（mutually constrained sequence）的数量不断增加，这表明非编码约束序列的演化周转速度极快。我们估算，当前约一半的非编码约束序列在过去约1.3亿年中发生了获得或丢失（以分化时间为单位的半衰期为d₁/₂ = 0.25–0.31）。尽管我们识别的短寿命约束序列富集了ENCODE生化注释信息，但诸多这类序列无法被针对更广范围泛哺乳动物保守性优化的模型所检测到。受约束的DNase I超敏感位点（DNase 1 hypersensitivity sites）、启动子（promoters）与非翻译区（untranslated regions）的演化稳定性均高于长非编码RNA（long noncoding RNA）基因座，后者的演化周转速度尤其快。与之形成对比的是，蛋白质编码序列（protein coding sequence）的稳定性极强，估算其半衰期超过10亿年（d₁/₂ = 2.1–5.0）。通过外推法我们估算，当前人类基因组中有8.2%（7.1%–9.2%）的序列处于负选择之下，因此大概率具备功能性；而在人类与小鼠分化后，仅有2.2%的序列在两者中均保持了约束状态。这些结果表明，人类基因组的演化历史极具动态性，尤其是其具备生物学功能的非编码序列部分。