A single-cell multiomic approach in planarians to understand regeneration.

用于了解涡虫再生的单细胞多组学方法。

• 批准号: MR/W017539/1
• 负责人: Jordi Solana
• 金额: $ 69.94万
• 依托单位: Oxford Brookes University
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FW017539%2F1
• 关键词: single; cell; multiomic; approach; planarians

Many animals are able to regenerate missing body parts and organs when they are injured. This ability, very reduced in humans and other mammals, is present in animals such as freshwater planarians. The planarian Schmidtea mediterranea can regenerate any body part in a matter of days. Thanks to its amenability for experimentation and ease of research use, S. mediterranea has become a powerful laboratory model where to study regeneration. Understanding this process is fundamental for regenerative medicine, as it seeks to recreate it with human cells in the laboratory, and ultimately in the clinic.Planarians regenerate thanks to a stem cell population present in their adult stages, the so-called neoblasts. Years of research have revealed genes, signals, molecules and processes that are important for planarian regeneration. However, these studies have suffered from the lack of cellular resolution: we do not know in which cell types these signals are active and how the different cell types coordinate during the regeneration process to form a tissue as complex as a head or a tail. Partly this is because current techniques to dissect the process genetically typically blend all tissues and cell types into bulk samples. Other techniques with high cellular resolution suffer from low throughput, giving information of a few genes in each assay.We have recently helped develop and used a very novel set of techniques that are revolutionising biology: single-cell analysis. It allows profiling the genetic information of thousands of individual cells by sequencing messenger RNAs and gene regulatory elements from each of them. With this, we can classify the major planarian cell types such as muscle, neurons, epidermis and dozens more and reconstruct their developmental processes. These methods have also been used in other developmental systems such as frogs, fish and newts, leading the way to a single-cell revolution of stem cell and developmental biology. The latest version of this method that we have already set up allow us to profile ~10-100 times more cells per experiment and makes feasible performing large sample sets with replicate experiments. This is a key modification to obtain statistically significant information.Thus, to understand how planarian regeneration works at the level of the individual cells that participate in the process, we will obtain single cell analysis profiles of around half a million cells, from different regeneration times, body parts and in replicates. Analysing all this information will allow us to elucidate which genes are activated in each cell type and at what time points during the process. We will perturb the function of these genes using well-established planarian methods to obtain functional information about the role of these genes from the effects of the perturbation. We will analyse these by further single-cell transcriptomic experiments. All of these experiments will tell us how each individual cell type behaves in the regeneration process. With such technique and cell numbers, even the rarest cell types will be captured. We will elucidate the signals they use to instruct the regenerative cells to integrate in the new regenerating tissue, and the functional effects of these signals.Altogether, these experiments will allow us to analyse planarian regeneration in an unprecedented quantitative way and at the single-cell level and with functional information. This will be a significant step towards understanding animal regeneration. Our data will spur novel studies on planarians to study the regulators and processes that we uncover and lead the way in the study of animal regeneration in other animals. The tools and methods that we will use for the first time will become standard in the study of regeneration and developmental processes, and key for single-cell biology. Understanding how animals regenerate is key to advance the agenda of human regenerative medicine.

许多动物在受伤时能够再生缺失的身体部位和器官。这种能力，在人类和其他哺乳动物中非常减少，存在于淡水真涡虫等动物中。扁涡虫Schmidtea medialacea可以在几天内再生身体的任何部位。由于其易于实验和易于研究使用，S。地中海已经成为研究再生的强大实验室模型。了解这一过程是再生医学的基础，因为它试图在实验室中用人类细胞重现这一过程，并最终在临床上使用。Planarians再生得益于成体阶段存在的干细胞群，即所谓的neoblast。多年的研究已经揭示了基因，信号，分子和过程，是重要的涡虫再生。然而，这些研究缺乏细胞分辨率：我们不知道这些信号在哪些细胞类型中是活跃的，以及不同的细胞类型在再生过程中如何协调形成像头部或尾部一样复杂的组织。部分原因是因为目前的技术，解剖过程中，基因通常混合所有组织和细胞类型到散装样品。其他高细胞分辨率的技术存在通量低的问题，每次检测只能提供少数基因的信息。我们最近帮助开发并使用了一套非常新颖的技术，这是生物学的革命：单细胞分析。它允许通过对每个细胞的信使RNA和基因调控元件进行测序来分析数千个细胞的遗传信息。有了这个，我们可以分类主要的涡虫细胞类型，如肌肉，神经元，表皮和几十个更多，并重建其发育过程。这些方法也被用于其他发育系统，如青蛙，鱼类和蝾螈，导致干细胞和发育生物学的单细胞革命。我们已经建立的这种方法的最新版本允许我们在每个实验中分析约10-100倍的细胞，并使重复实验的大样本集变得可行。这是一个关键的修改，以获得统计上有意义的信息。因此，为了了解在参与该过程的单个细胞水平上，涡虫再生是如何工作的，我们将获得来自不同再生时间、身体部位和重复的约50万个细胞的单细胞分析谱。分析所有这些信息将使我们能够阐明哪些基因在每种细胞类型中被激活，以及在这个过程中的什么时间点被激活。我们将扰动这些基因的功能，使用完善的涡虫方法，以获得功能信息的作用，这些基因的扰动的影响。我们将通过进一步的单细胞转录组学实验来分析这些。所有这些实验将告诉我们每种细胞类型在再生过程中的行为。有了这样的技术和细胞数量，即使是最稀有的细胞类型也会被捕获。我们将阐明它们用来指导再生细胞整合到新的再生组织中的信号，以及这些信号的功能效应。总之，这些实验将使我们能够以前所未有的定量方式，在单细胞水平和功能信息上分析涡虫再生。这将是理解动物再生的重要一步。我们的数据将刺激对涡虫的新研究，研究我们发现的调节因子和过程，并在其他动物的动物再生研究中发挥主导作用。我们将首次使用的工具和方法将成为再生和发育过程研究的标准，也是单细胞生物学的关键。了解动物如何再生是推进人类再生医学议程的关键。

项目成果

Single-cell transcriptomics in planaria: new tools allow new insights into cellular and evolutionary features.

• DOI: 10.1042/bst20210825
• 发表时间: 2022-10-31
• 期刊: Biochemical Society transactions
• 影响因子: 3.9

Allometry of cell types in planarians by single cell transcriptomics

• DOI: 10.1101/2023.11.01.565140
• 发表时间: 2023-11
• 期刊: bioRxiv
• 作者: Elena Emili;Alberto Pérez-Posada;Marika Christodoulou;Jordi Solana