The Tunicate Ciona: A New Model for the Effects of Aging on Tissue Regeneration

摘要 衰老过程中再生能力的丧失是生物学和生物医学领域中一个尚未解决的重大问题。 在这里，我们继续开发被囊状肠衣作为再生衰老研究的模型。乔纳 有许多有利的属性用于这一目的，包括一个小的测序基因组，一个广泛的工具包 基因操作，短暂的寿命，随着年龄增长而消失的强大的再生能力，以及 参与再生的干细胞生态位--一组稳定的绿色荧光蛋白转基因株系 在特定组织中表达，以及产生包含年轻和老年身体部位的嵌合动物的能力。 最重要的是，乔纳属于一种无脊椎动物脊索动物，被认为是与它最接近的近亲 脊椎动物，这表明对这种有机体的研究通常与脊椎动物有关，包括 人类。这项提案的三个具体目标旨在逐步加深我们的理解 干细胞巢、胚泡和周围组织环境在再生中的相对作用 衰老及其调控这一过程的分子机制--以芝麻纳口腔虹吸(口)为模型。 第一个目标将使用一种新的嵌合体试验，在这种试验中，干细胞的壁龛在年轻人和老年人之间交换。 将动物和分离的干细胞注射到老年动物体内，以确定干细胞的相对作用 生态位、干细胞本身、再生胚泡和周围组织环境 再生衰老。这一目标将利用我们之前对鳃囊(咽)干细胞的鉴定。 作为祖细胞来源的利基细胞迁移虹吸截肢部位并分化为 再生过程中的组织。第二个目标是识别、表征和本地化键的表达 负责幼年动物虹吸再生和丧失虹吸能力的调控基因 在古老的动物身上。这一目标将使用一组预先存在的候选基因，在 幼年动物的再生虹吸以及一组新的下调和上调的调控基因 在成体生命不同阶段通过转录组测序鉴定的干细胞利基中表达 周而复始。第三个目标是确定功能、等级关系和恢复丢失的能力 RNAi介导的基因敲除和改良检测关键再生基因的再生能力 在老动物身上使用嵌合体方法。这项研究旨在揭示遗传机制。 无脊椎动物脊索动物模型衰老过程中再生能力丧失的潜在原因 阐明我们对脊椎动物进化史和再生衰老原因的理解。

Summary The loss of regenerative capacity during aging is a major unsolved problem in biology and biomedicine. Here we continue to develop the tunicate Ciona intestinalis as a model for regenerative aging research. Ciona has many favorable attributes for this purpose, including a small sequenced genome, an extensive tool kit for genetic manipulations, a short life span, powerful regeneration capacities that disappear with age, an identified stem cell niche involved in regeneration, a collection of stable transgenic lines with green fluorescent protein expressed in specific tissues, and the ability to produce chimeric animals containing young and old body parts. Most importantly, Ciona belongs to an invertebrate chordate group recognized as the closest living relative of vertebrates, suggesting that research on this organism will be generally relevant to vertebrates, including humans. The three specific aims of this proposal are designed to incrementally advance our understanding the relative roles of the stem cell niche, the blastema, and the surrounding tissue environment in regenerative aging and the molecular mechanisms governing this process using the Ciona oral siphon (mouth) as a model. The first aim will use a novel chimera assay in which stem cell niches are swapped between young and old animals and isolated stem cells are injected into old animals to determine the relative roles of the stem cell niche, the stem cells themselves, the regeneration blastema, and the surrounding tissue environment in regenerative aging. This aim will exploit our previous identification of the branchial sac (pharynx) stem cell niche as the source of progenitor cells that migrate the site of siphon amputation and differentiate into multiple tissues during regeneration. The second aim is to identify, characterize, and localize the expression of key regulatory genes that are responsible for robust siphon regeneration in young animals and loss of this capacity in old animals. This aim will use both a set of pre-existing candidate genes expressed at high levels in the regenerating siphon of young animals as well as a new set of downregulated and upregulated regulatory genes expressed in the stem cell niche to be identified by transcriptome sequencing at different stages of the adult life cycle. The third aim is to determine the functions, hierarchical relationships, and ability to restore lost regenerative capacity of key regeneration genes by RNAi mediated gene knockdown and amelioration assays using the chimera approach in old animals. This research is designed to reveal the genetic mechanisms underlying the loss of regenerative capacity during aging in an invertebrate chordate model with the potential to clarify our understanding of the evolutionary history and causes of regenerative aging in vertebrates.