Challenging the dogma: is PABP-mediated post-transcriptional control essential in mammals?

挑战教条：PABP 介导的转录后控制对于哺乳动物至关重要吗？

• 批准号: BB/V016911/1
• 负责人: Nicola Gray
• 金额: $ 61.99万
• 依托单位: University of Edinburgh
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2021
• 资助国家: 英国
• 起止时间: 2021 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FV016911%2F1
• 关键词: Challenging; dogma; PABP; mediated; post

The proteins that make up our body are encoded in DNA as genes that serve as a genetic blueprint. The information in genes is decoded to produce proteins by a multi-step process known as gene expression. In this process, the DNA is first copied into an mRNA template (transcription), which is used to make proteins (translation). Cells need to make the right proteins, at the right time, place and in the correct amount so they can function properly. This means that their gene expression has to be tightly regulated. When these control mechanisms break down it can lead to a wide variety of diseases including cancer, metabolic, neurological and reproductive disorders. Manipulating these regulatory mechanisms can also benefit industrial processes that require efficient synthesis of proteins, for instance the production of antibodies. Both steps of the gene expression pathway can be regulated. Regulation at the first step is known as transcriptional control whereas, regulation of the second is called post-transcriptional control. Post-transcriptional control is critical as it affects more than half of all human genes, and is achieved by special "regulatory" proteins known as RNA-binding proteins (RBPs) and human cells can express thousands of RBPs. One family of regulatory RBPs that have been extensively studied are the poly(A)-binding proteins (PABPs). Based mainly on studies of one member, PABP1, this family have been shown to be key regulators of gene expression which have many different functions. PABP1 is considered to be so important that it is thought to be needed in every cell of the body. However, most of this knowledge comes from "transformed" cells growing in culture media, and it is unlikely they accurately reflect the functions of different cells and tissues in the body. Remarkably, therefore, despite intensive study over several decades, we still don't know what the biological roles of mammalian PABP1 are. For instance, is it essential for development? Here we aim to address this crucial gap in our knowledge by creating a so-called a "knock-out" mouse, in which PABP1 has been removed from all cells of the body. This will determine what processes and tissues within the body PABP1 is important for (e.g. brain development). Contrary to the view it is essential everywhere, we propose PABP1 is only critical for certain developmental stages, cell types or states, dependent on a number of factors including the presence of other family members. Therefore, these mice may be able to complete development but are unlikely to be "normal", for instance, they may have heart or fertility problems. In the longer term this may help us understand the basis of these disorders. To explore the hypothesis that a second family member, PABP4, can normally compensate for some, but not all of PABP1 functions in particular cell types, we will directly test this by making the first "double knock-out" PABP mouse. Importantly this will determine whether cells and tissues can function without any PABPs to regulate their post-transcriptional gene expression. We do not expect these mice to be able to complete development. Knowing why they die, will tell us for the first time what these proteins are normally so important for in the body.This is "blue-sky discovery" science and the results are likely to raise many more new hypothesis and questions. Importantly, the mice that we generate here are a flexible and refined tool to tackle new questions and our expertise place us in an excellent position to exploit these future opportunities. As mice are a considered a good genetically accessible model for human disease, we envisage in the longer term that our results will be relevant to human lifelong health.

构成我们身体的蛋白质在DNA中编码为基因，作为遗传蓝图。基因中的信息被解码，通过一个多步骤的过程产生蛋白质，这个过程被称为基因表达。在这个过程中，DNA首先被复制到mRNA模板中（转录），mRNA模板用于制造蛋白质（翻译）。细胞需要在正确的时间、地点和正确的数量制造正确的蛋白质，这样它们才能正常工作。这意味着它们的基因表达必须受到严格调控。当这些控制机制崩溃时，它可能导致各种各样的疾病，包括癌症，代谢，神经和生殖障碍。操纵这些调节机制也可以使需要有效合成蛋白质的工业过程受益，例如抗体的生产。基因表达途径的两个步骤都可以被调节。第一步的调控被称为转录控制，而第二步的调控被称为转录后控制。转录后控制是至关重要的，因为它影响超过一半的人类基因，并通过称为RNA结合蛋白（RBP）的特殊“调节”蛋白实现，人类细胞可以表达数千种RBP。已被广泛研究的调节RBP的一个家族是聚（A）结合蛋白（PABP）。主要基于对PABP 1的研究，该家族已被证明是具有许多不同功能的基因表达的关键调节因子。PABP 1被认为是如此重要，以至于它被认为是身体的每个细胞都需要的。然而，这些知识大部分来自于在培养基中生长的“转化”细胞，它们不太可能准确地反映体内不同细胞和组织的功能。值得注意的是，尽管经过几十年的深入研究，我们仍然不知道哺乳动物PABP 1的生物学作用。例如，它对发展是否至关重要？在这里，我们的目标是通过创造一种所谓的“敲除”小鼠来解决我们知识中的这一关键空白，在这种小鼠中，PABP 1已经从身体的所有细胞中去除。这将决定PABP 1对身体内的哪些过程和组织是重要的（例如大脑发育）。与PABP在任何地方都是必不可少的观点相反，我们认为PABP 1仅对某些发育阶段、细胞类型或状态至关重要，这取决于许多因素，包括其他家族成员的存在。因此，这些小鼠可能能够完成发育，但不太可能“正常”，例如，它们可能有心脏或生育问题。从长远来看，这可能有助于我们了解这些疾病的基础。为了探索第二个家族成员PABP 4在特定细胞类型中通常可以补偿PABP 1的一些但不是全部功能的假设，我们将通过制造第一个“双敲除”PABP小鼠来直接测试这一点。重要的是，这将决定细胞和组织是否可以在没有任何PABP的情况下发挥作用来调节其转录后基因表达。我们不期望这些小鼠能够完成发育。知道它们为什么会死，将首次告诉我们这些蛋白质在体内通常是什么样的重要。这是“蓝天发现”科学，其结果可能会提出更多新的假设和问题。重要的是，我们在这里生成的小鼠是一种灵活而精致的工具，可以解决新问题，我们的专业知识使我们处于利用这些未来机会的绝佳位置。由于小鼠被认为是人类疾病的良好遗传可及模型，我们设想从长远来看，我们的结果将与人类终身健康相关。