Combining viral and ribosomal mRNA capture technologies to develop a versatile system for neuronal transcriptome profiling

研究特定神经元组的功能(S)是非常困难的，因为大脑包含数以千计的不同类型的细胞，这些细胞类型是并列和相互联系的，并且根据它们的大小、形状、投射和功能而不同。最近已经开发出一种方法，允许所有正在被翻译成蛋白质的分子(信使RNA)在特定的神经元群体中被定义。这是一个非常强大的工具，因为第一次可以识别控制神经功能的基因集(例如，控制记忆形成的基因)和那些随年龄变化的基因，以及人类神经退行性疾病和神经精神疾病的基因。然而，这种方法依赖于昂贵的转基因小鼠品系，而且每次提出新的科学问题时，都需要产生一个新的小鼠品系。这项技术本身是相当复杂和耗时的，因为需要开发和繁殖(在许多情况下还杂交)一个或多个小鼠品系。这些考虑严重限制了这项技术对研究人员的可用性，相反，必须结合使用功能较弱的方法。在这项研究中，我们正在结合武田和布里斯托尔大学的专业知识来开发两种新的方法。这些方法使用病毒载体而不是转基因小鼠(称为病毒陷阱)，并允许以以前仅用转基因小鼠无法实现的速度、精度和多功能性来描述神经元。这些病毒捕获方法也将导致研究人员使用更少的动物。布里斯托尔大学将使用病毒捕获技术来识别一种名为支架附着因子(SAF)调节的RNA结合蛋白家族的基因和蛋白质。特别是，这些信息将被用来理解SAF蛋白如何管理调节记忆形成和衰老的过程。病毒捕捉技术将使武田能够进行详细的分析实验，而不需要冗长、昂贵和动物密集型的转基因程序。重要的是，由于病毒捕获技术可以移植到武田目前使用的其他模型，这项技术将提供一个新的平台，极大地促进基础生物学和未来的药理反应研究。最终，武田的目标是利用这项技术来进一步了解神经元基因调控和动态平衡以应对挑战，并利用这些洞察力来确定中枢神经系统紊乱的新靶点。

It is very difficult to study the function(s) of specific groups of neurons because the brain contains thousands of different cell types that are juxtaposed and interconnected and vary according to their size, shape, projections and function. Recently a method has been developed that allows all the molecules (messenger RNA) that are being translated into proteins to be in defined in specific neuronal populations. This is a very powerful tool as for the first time the sets of genes that are governing neuronal function (e.g. those controlling the formation of memories) and those that are altered with age and in human neurodegenerative and neuropsychiatric illnesses can be identified. However, this method is dependent on costly transgenic mice lines, and each time a new scientific question is asked a new mouse line needs to be generated. The technique itself is quite complex, and time consuming due to the need to develop and breed (and in many cases cross breed) one or more mouse lines. These considerations severely limit the availability of this technique to researchers and instead a combination of less powerful approaches must be used. In this study we are combining the expertise of Takeda and the University of Bristol to develop two new methods. These methods use viral vectors instead of transgenic mice (called viral TRAP) and allow neurons to be profiled with a speed, precision and versatility not previously possible with transgenic mice alone. These viral TRAP methods will also result in fewer animals being used by researchers. The viral TRAP technique will be used by the University of Bristol to identify the genes and proteins a family of RNA binding proteins called scaffold attachment factors (SAF) regulate. In particular this information will be used to understand how SAF proteins govern the processes that regulate memory formation and ageing. The viral TRAP technology will enable Takeda to perform detailed profiling experiments without the need for lengthy, expensive and animal-intensive transgenic programs. Importantly as the viral TRAP technique is portable to other models that Takeda currently use the technology will provide a novel platform that will greatly facilitate both basic biology and future pharmacological response studies. Ultimately, the objective of Takeda is to use this technology to further the understanding of neuronal gene regulation and homeostasis in response to challenge, and to use these insights to identify novel targets for central nervous system disorders.