The functional and evolutionary impacts of human-specific deletions in conserved elements.
The functional and evolutionary impacts of human-specific deletions in conserved elements.
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DOI：
10.1126/science.abn2253
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发表时间：
2023-04-28
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影响因子：
--
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破译将人类与灵长类近亲区分开来的分子和遗传变化对于了解我们的起源至关重要。尽管早期的研究优先考虑了新获得的基因序列或变异如何促进进化创新，但序列丢失的作用却很少受到重视。生物功能丰富的进化保守区域的改变尤其更有可能产生表型效应。因此，我们试图识别和表征在进化过程中保守的序列，但随后令人惊讶地在所有人类中丢失了。这些人类特有的保守区缺失（hCONDEL）可能在人类独特性状中发挥重要作用。测序技术的进步已经确定了黑猩猩和人类基因组之间数百万个遗传变化；然而，我们物种之间约 1% 至 5% 的差异对功能的影响在很大程度上尚不清楚。 hCONDEL 是这些主要非编码序列变化中的一类。尽管先前已经鉴定出大的 hCONDEL (>1 kb)，但绝大多数 hCONDEL (95.7%) 都很小（<20 个碱基对），并且尚未进行功能评估。我们采用大规模并行报告基因检测 (MPRA) 来表征数千个此类小型 hCONDEL 的效应，并发现了数百个具有功能效应的结果。通过了解这些 hCONDEL 的作用，我们可以深入了解驱动人类基因组进化的机制模式。我们通过检查不同脊椎动物基因组中的保守区域并与可靠注释的人类特异性固定缺失重叠，确定了 10,032 个 hCONDEL。我们发现这些 hCONDEL 被富集以删除源自干羊膜动物的保守序列。与转录、表观基因组和表型数据集的重叠都暗示了神经元和认知功能的影响。我们使用 MPRA 在六种不同的人类细胞类型中对这些 hCONDEL 进行了表征，包括诱导多能干细胞衍生的神经祖细胞。我们发现 800 个 hCONDEL 显示出物种特异性的调节效应。尽管许多 hCONDEL 会扰乱活性增强子中的转录因子结合位点，但我们估计 30% 的 hCONDEL 会创建或改善结合位点，包括激活子和阻遏子。一些 hCONDEL 表现出影响核心神经发育基因的分子功能。一个hCONDEL删除了神经发生基因HDAC5的活性增强子中的一个碱基，另一个hCONDEL删除了PPP2CA（一种调节神经元信号传导的基因）的替代启动子中的六个碱基。我们深入表征了 LOXL2（一种控制神经元分化的基因）的假定调控元件中的 hCONDEL。利用基因组工程将保守的黑猩猩序列重新引入人类细胞，我们证实人类缺失改变了 LOXL2 的转录输出。这些细胞的单细胞 RNA 测序揭示了 hCONDEL 诱导的一系列髓鞘形成和突触功能相关的转录变化。我们鉴定了数百个具有功能影响的 hCONDEL，揭示了可能塑造了我们独特的生物谱系的新分子变化。这些 hCONDEL 在多种生物系统中显示出预测的功能，但在神经元组织中的功能尤其丰富。许多 hCONDEL 诱导了调节活性的增强，这是一个令人惊讶的发现，因为人们通常认为删除保守碱基会消除功能。我们的工作为塑造人类或其他动物物种特异性生物学的核苷酸变化的表征提供了一个范例。人类特异性缺失，从其他动物高度保守的区域中去除核苷酸（hCONDEL）。我们评估了不同的生物学相关数据集中的 10,032 个 hCONDEL，并确定了组织特异性富集（左上）。通过比较 MPRA 中的黑猩猩和人类序列来表征 hCONDEL 的调节影响（左下）。评估了 hCONDEL 改善或干扰激活和抑制基因调控元件的能力（右上）。删除的黑猩猩序列被重新引入人类细胞，导致调节 LOXL2 的 hCONDEL 出现一系列转录差异（右下）。人类保守基因组序列被破坏可能是人类独特表型特征的基础。我们鉴定并表征了 10,032 个人类特异性保守缺失 (hCONDEL)。这些短的（平均 2.56 个碱基对）缺失丰富了遗传、表观基因组和转录组数据集中的人脑功能。通过对六种细胞类型进行大规模平行报告分析，我们发现 800 个 hCONDEL 在调节活性方面具有显着差异，其中一半增强而不是破坏调节功能。我们重点介绍了几种对大脑发育具有假定的人类特异性影响的 hCONDEL，包括 HDAC5、CPEB4 和 PPP2CA。将 hCONDEL 恢复为祖先序列会改变 LOXL2 的表达以及参与髓鞘形成和突触功能的发育基因。我们的数据为研究驱动人类和其他物种新特征的进化机制提供了丰富的资源。
Deciphering the molecular and genetic changes that differentiate humans from our closest primate relatives is critical for understanding our origins. Although earlier studies have prioritized how newly gained genetic sequences or variations have contributed to evolutionary innovation, the role of sequence loss has been less appreciated. Alterations in evolutionary conserved regions that are enriched for biological function could be particularly more likely to have phenotypic effects. We thus sought to identify and characterize sequences that have been conserved across evolution, but are then surprisingly lost in all humans. These human-specific deletions in conserved regions (hCONDELs) may play an important role in uniquely human traits. Sequencing advancements have identified millions of genetic changes between chimpanzee and human genomes; however, the functional impacts of the ~1 to 5% difference between our species is largely unknown. hCONDELs are one class of these predominantly noncoding sequence changes. Although large hCONDELs (>1 kb) have been previously identified, the vast majority of all hCONDELs (95.7%) are small (<20 base pairs) and have not yet been functionally assessed. We adapted massively parallel reporter assays (MPRAs) to characterize the effects of thousands of these small hCONDELs and uncovered hundreds with functional effects. By understanding the effects of these hCONDELs, we can gain insight into the mechanistic patterns driving evolution in the human genome. We identified 10,032 hCONDELs by examining conserved regions across diverse vertebrate genomes and overlapping with confidently annotated, human-specific fixed deletions. We found that these hCONDELs are enriched to delete conserved sequences originating from stem amniotes. Overlap with transcriptional, epigenomic, and phenotypic datasets all implicate neuronal and cognitive functional impacts. We characterized these hCONDELs using MPRA in six different human cell types, including induced pluripotent stem cell–derived neural progenitor cells. We found that 800 hCONDELs displayed species-specific regulatory effect effects. Although many hCONDELs perturb transcription factor–binding sites in active enhancers, we estimate that 30% create or improve binding sites, including activators and repressors. Some hCONDELs exhibit molecular functions that affect core neurodevelopmental genes. One hCONDEL removes a single base in an active enhancer in the neurogenesis gene HDAC5, and another deletes six bases in an alternative promoter of PPP2CA, a gene that regulates neuronal signaling. We deeply characterized an hCONDEL in a putative regulatory element of LOXL2, a gene that controls neuronal differentiation. Using genome engineering to reintroduce the conserved chimpanzee sequence into human cells, we confirmed that the human deletion alters transcriptional output of LOXL2. Single-cell RNA sequencing of these cells uncovered a cascade of myelination and synaptic function–related transcriptional changes induced by the hCONDEL. Our identification of hundreds of hCONDELs with functional impacts reveals new molecular changes that may have shaped our unique biological lineage. These hCONDELs display predicted functions in a variety of biological systems but are especially enriched for function in neuronal tissue. Many hCONDELs induced gains of regulatory activity, a surprising discovery given that deletions of conserved bases are commonly thought to abrogate function. Our work provides a paradigm for the characterization of nucleotide changes shaping species-specific biology across humans or other animals. Human-specific deletions that remove nucleotides from regions highly conserved in other animals (hCONDELs). We assessed 10,032 hCONDELs across diverse, biologically relevant datasets and identified tissue-specific enrichment (top left). The regulatory impact of hCONDELs was characterized by comparing chimp and human sequences in MPRAs (bottom left). The ability of hCONDELs to either improve or perturb activating and repressing gene-regulatory elements was assessed (top right). The deleted chimpanzee sequence was reintroduced back into human cells, causing a cascade of transcriptional differences for an hCONDEL regulating LOXL2 (bottom right). Conserved genomic sequences disrupted in humans may underlie uniquely human phenotypic traits. We identified and characterized 10,032 human-specific conserved deletions (hCONDELs). These short (average 2.56 base pairs) deletions are enriched for human brain functions across genetic, epigenomic, and transcriptomic datasets. Using massively parallel reporter assays in six cell types, we discovered 800 hCONDELs conferring significant differences in regulatory activity, half of which enhance rather than disrupt regulatory function. We highlight several hCONDELs with putative human-specific effects on brain development, including HDAC5, CPEB4, and PPP2CA. Reverting an hCONDEL to the ancestral sequence alters the expression of LOXL2 and developmental genes involved in myelination and synaptic function. Our data provide a rich resource to investigate the evolutionary mechanisms driving new traits in humans and other species.
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DOI：
10.1093/bioinformatics/btr064
发表时间：
2011-04-01
期刊：
Bioinformatics (Oxford, England)
影响因子：
--
作者：
Grant CE;Bailey TL;Noble WS
通讯作者：
Noble WS
影响因子：
4
作者：
Dennis MY;Eichler EE
通讯作者：
Eichler EE
影响因子：
4
作者：
Bolós, V;Peinado, H;Cano, A
通讯作者：
Cano, A
影响因子：
64.5
作者：
Griesemer D;Xue JR;Reilly SK;Ulirsch JC;Kukreja K;Davis JR;Kanai M;Yang DK;Butts JC;Guney MH;Luban J;Montgomery SB;Finucane HK;Novina CD;Tewhey R;Sabeti PC
通讯作者：
Sabeti PC
DOI：
10.1073/pnas.1932072100
发表时间：
2003-09-30
影响因子：
11.1
作者：
Kent, WJ;Baertsch, R;Haussler, D
通讯作者：
Haussler, D