科研新进展：小鼠大脑普遍存在细胞嵌合

//原作者Dylan Z.Faltine-Gonzalez和Justus M. Kebschull2022年9月发表于Science，原标题：A mosaic of new and old cell type

//翻译 小鹰

（严禁用于商业用途）

Comparative transcriptomics could reveal patterns of cell type evolution in the tetrapodBrain

比较转录组学（译者注：比较转录组学注重基因序列层面的变化，研究内容多为进化学上所谓系统发育关系，特别是因经济或技术问题不宜用于基因组学）可以揭示不同类型细胞在四足动物（译者注：不严谨的讲，两栖纲，爬行纲，哺乳纲为四足动物）脑部的进化

Over the past decade, hundreds of cell types have been identified in specialized brain regions of the laboratory mouse. How this staggering persity of cell types and regions evolved is currently unknown.

在过去的十年里，在实验室小鼠的特定脑区已经鉴定出数百种不同类型的细胞。这种惊人的细胞类型和细胞区域的多样性是如何进化的，目前尚不清楚。

On pages 1060, 1063, 1061, and 1062 of this issue, Hain et al (1), Woych et al (2), Lust et al (3), and Wei et al (4), respectively, leverage singlecell and spatial transcriptomics in reptiles and amphibians to investigate cell type evolution at the brain scale. Hain et al produce a whole-brain cell atlas of the bearded dragon. Woych et al profile the developing and adult salamander telencephalon.

在本期杂志的第1060、1063、1061和1062页上，Hain等人(1)、Woych等人(2)、Lust等人(3)和Wei等人(4)分别利用爬行动物和两栖动物的单细胞和空间转录组学（译者注：空间转录组学可以同时获得细胞的空间位置信息和基因表达数据，对组织原位细胞真实基因表达进行研究。现有的空间转录组技术主要分为两类：一类是基于杂交和成像的方法，例如 FISH；另一类则是基于测序的方法）来研究大脑层次的细胞类型进化。Woych等人制作了一份胡须蜥（译者注：似乎确实资料）的全脑细胞图谱。Woych等人描述了亚成体的和成体火蜥蜴的端脑（译者注：由前脑发育形成，不严谨的讲，端脑和大脑（皮层）同源）。

Lust et al and Wei et al tackle the axolotl telencephalon during development and regeneration using complementary single-cell multi-omic and new spatial transcriptomic techniques. Together, these studies reveal that rather than being a set of old and new regions, vertebrate brains are formed from a mosaic of conserved and new cell types.

Lust等人和Wei等人利用互补的单细胞多组学技术（译者注：多组学技术包括基因组学、转录组学、蛋白质组学、代谢组学、影像组学等）和新的空间转录技术解决了钝口螈（译者注：两栖纲有尾目 学名为A.mexicanum）端脑在发育和再生过程中的问题。总而言之，这些研究表明，与其说是新旧两组区域，不如说是原始和新生细胞类型的嵌合。

Traditionally, brain evolution is studied by identifying homologous brain regions across disparate species on the basis of cytoarchitecture, marker gene expression, and developmental origin. Thus, the brain is often considered a collection of “old” and “new” brain regions. The traditional approaches for region comparison do not resolve cell types. Nevertheless, region level conservation might generalize to the cell type level.

传统上，脑进化是通过根据细胞结构、标记基因表达和发育起源识别不同物种的同源脑区来研究的。（译者注：因为在胚胎发育及进化上的同源性，我们一般可将脊椎动物的大脑统一分类，如原脑皮（海马体），古脑皮（梨状叶），纹状体（基底核）等）因此，大脑通常被认为是“旧的”和“新的”大脑区域的集合。传统的区域比较方法不能区分细胞类型。然而，区域层面的现状可能推广到细胞类型层面。

Recently, a host of single-cell omics methods have been developed that query gene expression of dissociated single cells and can be applied to any species with an annotated transcriptome or, preferably, an annotated genome. Emerging spatial transcriptomic technologies are also now allowing the interrogation of many genes with high spatial resolution in tissue. These technologies provide the opportunity to systematically compare transcriptomic information across species. Pioneering comparative transcriptomic studies have begun to investigate brain cell type and region evolution in vertebrates outside of the mammalian lineage (5–8), but they have largely focused on inpidual regions.

现今，我们已经开发了许多单细胞组学方法来查证独立单细胞的基因表达，并且可以应用于任何具有注释的转录组（译者注：基因注释是指对基因功能的描述），更优的是，具有注释的基因组的物种。新兴的空间转录组学现在也允许以高空间分辨率研究组织中的许多基因。这些技术为系统地比较不同物种的转录信息提供了可能。开创性的比较转录组学研究已经开始用于研究哺乳动物以外脊椎动物的脑细胞类型和区域进化。但他们主要集中在个别领域。

Hain et al tested the old versus new brain region and cell type hypothesis by producing a cell type atlas of the brain of the bearded dragon Pogona vitticeps, a lizard. When comparing lizard and mouse data, they found that cells from broadly defined brain regions in both species correspond to each other, indicating conserved region-specific gene expression signatures, as expected from traditional methods.

Hain等人通过制作一种蜥蜴的大脑细胞类型图谱，验证了旧的和新的大脑区域和细胞类型假说。当比较蜥蜴和老鼠的数据时，他们发现两个物种来自广义脑区的细胞相互对应，表明有着保守的区域特异性基因表达的标志，正如传统方法所预期的那样。

These signatures might originate from developmental constraints that are preserved in conserved homeobox transcription factor expression patterns. However, when mapping cell types at higher resolution, the authors observed both similar and very dissimilar cell types across species in almost every brain pision investigated (telencephalon, diencephalon, mesencephalon), indicating the intermingling of both highly conserved and species-specific cell types.

这些标志可能起源于发育限制，这些发育限制保存在保守的同源框转录因子表达模式中。然而，当以更高的分辨率绘制细胞类型图时，作者在几乎每个所研究的脑区(端脑、间脑、中脑)中均观察到跨物种性的，相似和相反的细胞类型，表明高度保守的细胞类型和物种特有的细胞类型混合在一起。

Although in the telencephalon mostly inhibitory interneurons are conserved, in hypothalamus, tectum, and thalamus excitatory and inhibitory projection and local neurons show evidence of conservation. The existence of conserved and new cell types within conserved brain regions suggests that neuron types are evolutionarily plastic and capable of independently evolving new gene expression signatures and functions within their developmental framework.

虽然在端脑，大多数抑制性中间神经元是保守的，但在下丘脑、顶盖（译者注：又被译作四叠体，位于中脑背侧部，由两对圆形小丘组成，分别称为上丘）和下丘）和丘脑，兴奋性和抑制性投射和局部神经元显示出保守性。在保守的脑区中存在保守的和新生的细胞类型，这表明神经元类型在进化上是具有可塑性的，并能够在其发育进程架构内独立地进化新的基因表达特征和功能。

These findings resonate with and are extended by the studies of Lust et al, Wei et al, and Woych et al, which focus on the amphibian telencephalon, the part of the brain that in mammals contains the neocortex.

这些发现与Lust等人、Wei等人和Woych等人的研究产生了共鸣，并得到了扩展，他们专注于两栖动物的端脑，哺乳动物大脑中包含新脑皮的部分（译者注：在大脑中，不严谨地讲，除去海马体，纹状体，梨状叶的部分叫做新脑皮。但一些皮层下的脑组织，如基底神经节，嗅球和海马体也是由端脑发育形成的。）。

They reveal deeply conserved classes of telencephalic inhibitory cells from each of the three developmental origins that have been recognized in mammals. Similar to the findings of Hain et al in bearded dragons and previous findings in turtles (6), conservation of excitatory neurons in the telencephalon is generally lower, suggesting the evolution of new cell types. Nevertheless, broad similarities in gene expression allow both Woych et al and Lust et al to match pallial regions in the amphibian brain to their homologs in reptiles, birds, and mammals.

他们揭示了在哺乳动物中已被认知的三种发育来源中的每一种都具有高度保守的端脑抑制细胞类型。与Hain等人在胡须蜥上的发现以及之前在海龟上的发现相似(6)，端脑中兴奋性神经元的保守性通常较低，这支持新兴细胞类型的衍生。然而，基因表达的广泛相似性使Woych等人和Lust等人都能够将两栖动物大脑中的Pallial区域（译者注：Pallial似乎是大脑皮质（cerebral cortex）的意思）与它们在爬行动物、鸟类动物和哺乳动物中的同源区域相匹配。

The intermingling of conserved and new cell types within broad regions challenges the idea of clear-cut old and new regions at the cell type level. Nevertheless, evidence for evolutionary innovation that creates new regions with new cell types and functions is abundant. Woych et al find that salamander (Pleurodeles waltl) ventral pallium neurons are homologous to parts of the reptile dorsal ventricular ridge, but no homolog to the excitatory cells of another part of the same structure exists in the salamander.

在广泛的程度内，保守的和新生的细胞类型混合在一起，挑战了在细胞类型水平上明确划分新旧区域的想法。但是，有不少证据表明，进化创新创造了具有新细胞类型和功能的新区域。Woych等人发现P.Waltl大脑皮层腹侧神经元与爬行动物背侧脑室脊的部分神经元同源，但在相同结构中，另一部分的兴奋细胞不存在同源细胞。

The mammalian neocortex has no direct match at the level of excitatory cell types in the amphibian. However, cells matching mouse layer 4 neocortical cells seem to emerge with the reptiles. Similarly, Hain et al find evidence of region formation in the amniote thalamus, where medial thalamus appears well conserved between lizard and mouse as is a general mediolateral axis, but specific nuclei and their complement of cell types appear to be evolutionary innovations.

哺乳动物的新脑皮在两栖动物的兴奋细胞类型水平上没有直接匹配。然而，与小鼠第四层新皮层细胞相匹配的细胞似乎是爬行动物一起出现。同样，Hain等人发现了羊膜动物丘脑区域形成的证据。，内侧丘脑在蜥蜴和老鼠中似乎保存得很好，就像一般的丘脑内线核群一样，但特定的核群及它们的细胞类型的补充似乎是进化上的创新。

Additionally, the studies of Lust et al, Wei et al, and Woych et al investigate the developmental trajectories of brain regions and cell types observed in the adult. Woych et al note the conservation of transcription factor programs regionally specific to mouse pallium. Similarly, Lust et al identify the gene-regulatory networks underlying the regional persification of pallial excitatory neurons during postembryonic development in axolotls, and Wei et al examine spatial trajectories during axolotl development.

此外，Lust等人、Wei等人和Woych等人的研究调查了在成体中观察到的大脑区域和细胞类型的发育轨迹。Woych等人注意到小鼠大脑皮层特定区域转录因子程式的保守性。类似地，Lust等人发现了钝口螈胚胎后发育过程中胚层兴奋性神经元区域多样化的基础上的一般调控网络，Wei等人研究了钝口螈发育过程中的空间轨迹。

Moreover, Lust et al and Wei et al investigate the ability of axolotls to regenerate their telencephalon after injury. Both identified a distinct wound healing response to damage at the start of regeneration. Subsequently, proliferation and transition to neurogenesis seem to parallel that observed during development, leading to the reestablishment of all previously existing cell types and even longrange connections.

此外，Lust等人和Wei等人研究了钝口螈在损伤后再生端脑的能力。两人都确定了在再生开始时对损伤有明显的创口愈合反应。随后，增殖和向神经再生的转变似乎与发育过程中所观察到的相平行，导致所有先前存在的细胞类型以及长距离连接的重建。

The key to integrating the findings of intermingled old and new cell types and of evolutionary innovation likely rests on two concepts. Regions, like cell types, are thought to be hierarchically organized (9).Therefore, a coarsely defined old region might contain both old and new cell types.

整合新旧混合细胞类型和进化创新的发现的关键可能取决于两个理论。大脑分区就像细胞分型一样，被认为是按层次组织的(9)。因此，粗略定义的旧区域可能同时包含原始细胞类型和新兴细胞类型。

When investigated at a finer region resolution, however, these new and old cell types might spatially segregate into evolutionary newer and older subregions that evolved potentially by duplication and pergence of sets of cell types (8).

然而，当以更精细的区域分辨率进行分析时，这些新的和旧的细胞类型可能在空间上分离成衍生的新的和旧的亚区，这些亚区可能通过细胞类型集合的复制和分歧而进化(8)。

The second concept is that cell classes originating from different developmental niches can have different evolutionary paths and nevertheless intermingle in the adult (6–8). For example, well-conserved telencephalic interneurons migrate during development into pergent regions of the pallium, which results in the intermingling of old and new neurons in the adult. Like all evolutionary comparisons, comparative transcriptomics must distinguish orthology from convergent evolution.

第二个概念是，起源于不同发育环境的细胞类别可以有不同的演化路径，但在成体(6-8岁)中却嵌合在一起。例如，保存良好的端脑中间神经元在发育过程中迁移到大脑皮层的分歧区域，这导致成年后新旧神经元的混合。与所有进化比较一样，比较转录组学必须区分直系同源和趋同进化（译者注：趋同进化指源自不同祖先的生物，由于相似的生活方式，整体或部分形态结构向着同一方向改变，而直系同源指源自相同祖先的生物）。

Because transcriptomic space is high dimensional, this is a constant challenge. The studies of Hain et al, Woych et al, Lust et al, and Wei et al tackle this problem with a remarkable breadth of technology and species comparisons. In particular, Lust et al and Woych et al integrate transcriptomic data with spatial transcriptomics or whole-brain in situ hybridization, developmental data, and connectivity data.

因为转录空间是高维度的，这是一个持续的挑战。Hain等人、Woych等人、Lust等人和Wei等人的研究通过海量的技术和物种比较解决了这个问题。特别的，Lust等人和Woych等人将转录数据与空间转录组或全脑原位杂交、发育数据和连接数据相结合。

Wei et al then provide a glimpse into the future of high–spatial resolution comparative transcriptomics by using a new method called Stereo-seq, which allows transcriptome wide sequencing at subcellular resolution (10). These studies highlight the potential of applying the powerful transcriptomic methods that are usually reserved for mouse to nonstandard models (11). Each of the articles produced massive single-cell and often multimodal datasets and mined publicly available data, showcasing the importance of data sharing and the power of accumulating single-cell data from many species for evolutionary comparisons. They demonstrate that new and old cell types intermingle in broadly defined brain regions.

Wei等人然后通过使用一种名为Stereo-Seq（译者注：空间分辨的转录组技术）的新方法，对高空间分辨率比较转录学的未来进行了探索，这种方法允许在亚细胞分辨率下进行全转录组测序(10)。这些研究突显了将通常为小鼠保留的强大的转录组学应用于非标准模型的潜力(11)。每一篇文章都产生了大量的单细胞数据集，这往往是多模式数据集，并挖掘了公开可用的数据，展示了数据共享的重要性，以及积累来自众多物种的单细胞数据用于进化比较的力量。他们证明了新的和旧的细胞类型在广义的的大脑区域嵌合在一起。

Further studies with increased spatial resolution will be necessary to identify at what level of the cell type hierarchy and region hierarchy this model holds and how evolutionary innovations of brain regions and subregions interface with these findings. Understanding of evolutionary processes is needed both at a level that describes the adult phenotype, which should be most relevant for understanding brain function (12), and at a mechanistic and developmental level. j

随着空间分辨率的提高，将有必要进行进一步的研究，以确定该模型在细胞类型层次和区域层次的哪个水平上成立，以及大脑区级和亚区级的新生性演化如何与这些发现相结合。对进化过程的理解既需要在描述成体表型的水平上，这应该与理解大脑功能最相关(12)，也需要在机械和发育水平上。J