En1 sweat we trust: How the evolution of an Engrailed 1 enhancer made humans the sweatiest ape.
En1 sweat we trust: How the evolution of an Engrailed 1 enhancer made humans the sweatiest ape.
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我们信任的 En1 汗液：Engrailed 1 增强剂的进化如何使人类成为出汗最多的猿类。
DOI：
10.1080/23328940.2021.2019548
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发表时间：
2022
期刊：
影响因子：
--
通讯作者：
Kamberov,YanaG
中科院分区：
文献类型：
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作者：
Aldea,Daniel;Kamberov,YanaG
人类主要通过皮肤表面水分的蒸发来降温。人体的汗腺负责分泌蒸发冷却所需的水分，对这种体温调节机制至关重要。在灵长类动物中，人类的汗腺密度最高。事实上，人类的汗腺密度平均是黑猩猩和猕猴的10倍。人类皮肤中分泌腺的高密度是人类与其他灵长类动物最显著的表型差异之一，也是人类通过出汗降温的特殊能力的关键组成部分。尽管它很重要，但对这种适应性人类特征进化背后的遗传机制研究甚少。在我们最近的研究[3]中，我们确定了第一个负责人类内分泌腺进化的遗传和发育机制。我们的研究建立在我们和其他人之前的工作的基础上，表明转录因子En1 （En1）的上调促进了小鼠分泌腺的规范，En1表达的上调与人类分泌腺的发育相关[4,5]。因此，我们试图确定人类是否进化出了调控突变，导致EN1在皮肤中的表达增强，从而诱导更多汗腺的形成。由于En1在哺乳动物分泌腺形成区域的空间表达模式是保守的[4-6]，我们首先扫描了人类En1位点附近的非编码基因组，寻找可能作为该基因调控元件或增强子的进化保守序列。为此，我们利用比较基因组学结合转基因小鼠的功能测试来寻找在发育过程中表达EN1的皮肤细胞中显示活性的EN1候选增强子（ECEs）。我们确定了5种在小鼠小分泌形成皮肤中表现出这种发育促进活性的ECEs。其中一个阳性ECE的人类同源性与其他类人猿差异最大，我们将其命名为人类ECE18 （hECE18）。hECE18还值得注意的是，它与2xHAR20重叠，后者是一类计算定义的基因组元件之一，其特征是在非人类脊椎动物中具有高度序列保守性，但在人类中具有异常高的分化，被认为在产生人类特异性进化表型[7]中发挥作用。此外，我们发现与其他哺乳动物ECE18同源物相比，hee18在培养的人或小鼠皮肤细胞中具有最大的增强子活性。hee18的高活性是人类谱系中多个单核苷酸取代的累积的结果，因为与其他灵长类动物相比，其中两个人类突变与SP1蛋白具有高亲和力结合。这些结果促使我们在发育模型中测试hECE18增强子是否可以通过调节En1来影响分泌腺的数量。为此，我们使用CRISPR-Cas9基因组编辑技术将小鼠基因组中的内源性ECE18序列替换为同源的hECE18序列。值得注意的是，我们发现人类增强子上调En1在皮肤中的表达，以促进皮肤发育过程中更多分泌腺的规范。再加上我们在体外发现hee18活性增加，我们从发育小鼠模型中获得的数据与进化一致
Humans cool themselves primarily through the evaporation of water from the skin surface. Human eccrine sweat glands, which are responsible of secreting the water for evaporative cooling, are essential for this mechanism of thermoregulation [1]. Among primates, humans have the highest eccrine sweat gland density. Indeed, human eccrine sweat gland density is on average ten times that of chimpanzees and macaques [2]. The high density of eccrine glands in human skin is one of the most dramatic phenotypic differences that distinguishes humans from other primates, and is a key component that makes possible humans’ exceptional ability to cool off by sweating [1]. In spite of its importance, the genetic mechanisms behind the evolution of this adaptative human trait have been poorly studied. In our recent study [3], we identified the first genetic and developmental mechanism responsible for the evolutionary elaboration of human eccrine glands. Our study built on previous work from us and others showing that the upregulation of the transcription factor Engrailed 1 (En1) promotes the specification of eccrine glands in mice, and the upregulation of EN1 expression is correlated with eccrine gland development in humans [4, 5]. We therefore sought to determine if humans have evolved regulatory mutations that lead to a potentiation of EN1 expression in the skin to induce the formation of more eccrine glands. Since the spatial pattern of En1 expression is conserved across mammals in regions where eccrine glands form [4–6], we first scanned the non-coding genome near the human EN1 locus for evolutionarily conserved sequences with the potential to act as regulatory elements, or enhancers, of this gene. To this end we utilized comparative genomics coupled with functional testing in transgenic mice to look for EN1 candidate enhancers (ECEs) that showed activity in En1-expressing skin cells during development. We identified 5 ECEs that exhibited such developmental enhancer activity in mouse eccrine forming skin. The human homolog of one positive ECE, which we named human ECE18 (hECE18) was the most highly sequence diverged relative to other apes. hECE18 was also notable in that it overlapped 2xHAR20, one of a class of computationally-defined genomic elements characterized by high sequence conservation across non-human vertebrates but exceptionally high divergence in humans that have been postulated to play roles in generating human-specific evolutionary phenotypes [7]. In addition, we found that hECE18 has the greatest enhancer activity in cultured human or mouse skin cells as compared to other mammalian ECE18 homologs. The higher activity of hECE18 is the result of the accumulation of multiple single nucleotide substitutions that have specifically evolved on the human lineage, and because two of these human mutations give a high-affinity binding to the SP1 protein when compared to other primates.These results prompted us to test whether the hECE18 enhancer can affect the number of eccrine glands through the regulation of En1 in a developmental model. To that end we replaced the endogenous ECE18 sequence in the mouse genome with the homologous hECE18 sequence using CRISPR-Cas9 genome editing. Remarkably, we found that the human enhancer upregulates En1 expression in the skin to promote the specification of more eccrine glands during skin development. Coupled with our findings of increased hECE18 activity in vitro, our data from the developmental mouse model are consistent with an evolutionary