Epigenetic and signalling mechanisms underpinning impaired lymphocyte development and antibody recombination in ageing

该项目将采取自上而下和自下而上的方法。首先，我们将研究在已建立的小鼠模型中丢失这些成分的后果，在该模型中，相关基因在骨髓B细胞中特异性缺失。我们会问：缺失如何影响B细胞的发育进程、增殖、重组、抗体库的多样性？下游信号通路如何改变，这又如何影响转录组及其表观遗传驱动因子？其次，我们将研究这些成分是如何在表观遗传水平上进行调控的。我们的研究结果表明，它们被Polycomb介导的miRNA机制以及与远端增强子失去联系所抑制。我们将在体外B细胞系统中使用Crispr-Cas9靶向关键DNA元件和适当的抑制剂来研究这些机制，以确定衰老中受损的关键表观遗传调节因子。博士项目将包括实验（“湿”）和生物信息学（“干”）的组成部分。这些技术都是在实验室中完善的，包括流式细胞术、小鼠模型、信号传导测定、RNA-seq、VDJ-seq、捕获HiC、ChIP-seq、单细胞测序、生物信息学、Crispr-Cas敲除。Babraham生物信息学小组将提供生物信息学方面的广泛培训和援助。

This project will take a top-down and a bottom-up approach. First we will investigate the consequences of loss of these components in established mouse models in which the relevant genes are deleted specifically in bone marrow B cells. We will ask: How does deletion impact on B cell developmental progression, proliferation, recombination, diversity of the antibody repertoire? How are downstream signaling pathways altered and how does this in turn affect the transcriptome and its epigenetic drivers? Second, we will investigate how these components are regulated at the epigenetic level. Our findings suggest that they are repressed by Polycomb-mediated and miRNA mechanisms, and by loss of contact with distal enhancers. We will investigate these mechanisms using Crispr-Cas9 targetting of key DNA elements and appropriate inhibitors in in vitro B cell systems, to determine the key epigenetic regulators impaired in ageing. The PhD project will include both experimental ('wet') and bioinformatics ('dry) components. Techniques are all well-established in the lab and will include flow cytometry, mouse models, signaling assays, RNA-seq, VDJ-seq, capture HiC, ChIP-seq, single cell sequencing, bioinformatics, Crispr-Cas knockout. Extensive training and assistance in bioinformatics will be provided by the Babraham Bioinformatics Group.