Phase variable epigenetic control in firmicutes

一个物种的所有细胞都含有相同的DNA；区别它们的是DNA被激活（“转录”）的方式。调节DNA转录的一种方法是通过DNA本身的直接化学修饰，最常见的是通过甲基化过程。这被称为“表观遗传”调控。对人类和其他复杂生命形式的表观遗传调控进行了广泛的研究，但对细菌的研究相对较少。细菌基因组经常被广泛甲基化，这是“限制性修饰”（RM）系统的结果，该系统在特定位点修饰细胞DNA，使其与感染病毒的DNA区分开来。基因组中的许多甲基化位点都有可能实质性地影响基因的调节方式。然而，由于大多数RM系统是稳定的，因此不能作为一种监管机制。这不是我们最近在细菌肺炎链球菌中独立鉴定的两组基因的情况。肺炎球菌是一种共生细菌，通常由四分之一到三分之一的无症状幼儿携带，这是导致中耳感染、肺炎和脑膜炎等疾病的主要原因。我们发现的这组基因是RM系统，通过一组特定的DNA重排在数小时或数天内发生变化。这导致由RM系统引起的甲基化模式也在短时间尺度上发生变化。实验数据发现，不同形式的逆转录RM系统导致不同的甲基化模式，每种甲基化模式都与不同的基因表达模式相关。这种细菌基因的表观遗传调控被发现改变了细菌的毒力，一些甲基化模式使肺炎球菌更容易引起疾病。这可能是肺炎球菌从无害的共生菌转变为危险病原体的一个重要因素。本项目旨在通过研究肺炎球菌中的第二变量RM系统是否影响相同的过程，还是对细胞生理有不同的影响，以及该系统是否调节其他致病菌的毒力，来验证这一假设。对数千个公开的细菌DNA序列的搜索使我们能够识别出数百个拥有类似系统的物种。这些细菌包括在人类肠道中非常常见的细菌种类，一些存在于益生菌饮料中，另一些则与奶酪的生产有关。也许最重要的是，它们也存在于许多致病菌中。该项目旨在调查这些可变RM系统是否也可能调节三种细菌的毒力，每种细菌都代表对公众健康的主要威胁。第一种是猪链球菌，这是一种通常与猪有关的物种，它正在成为一种能够引起严重感染（如脑膜炎）的主要病原体。第二种是单核细胞增生李斯特菌，这是一种食源性细菌，可导致潜在的致命感染。第三种是粪肠球菌（Enterococcus faecalis），这是引起高度耐抗生素感染的主要原因，特别是在医院环境中。在这三个物种中，可变RM位点存在于与引起人类高水平疾病相关的谱系中，而在动物或人类中无症状发现的谱系中则不存在。该项目的总体目标是弄清楚这些系统如何在调节与细菌毒性有关的基因中发挥作用，以及它们是如何进化的以及它们的多样性。这些信息将使我们了解为什么这些不寻常的基因分布，以及为什么细菌从无害携带到引起疾病。这将更好地告知我们如何防止这种转变的策略，从而驯服这些常见但潜在危险的细菌。

All the cells of a species contain the same DNA; what distinguishes them is the way in which that DNA is activated ('transcribed'). One method of regulating DNA transcription is through direct chemical modification of the DNA itself, most commonly through a process of methylation. This is termed 'epigenetic' regulation. There has been extensive study of epigenetic regulation in humans and other complex life forms, but comparatively little in bacteria. Bacterial genomes are often extensively methylated, as a consequence of 'restriction modification' (RM) systems, which modify the cell's DNA at particular sites to allow it to be distinguished from the DNA of infecting viruses. The many sites of methylation across the genome have the potential to substantially affect the way in which genes are regulated. However, as most RM systems are stable and therefore cannot serve as a regulatory mechanism. This is not the case for two sets of genes we have recently independently characterised in the bacterium Streptococcus pneumoniae, the pneumococcus. The pneumococcus is a commensal bacterium, typically carried by between a quarter and a third of young children asymptomatically, that is a major cause of diseases including middle ear infections, pneumonia and meningitis. The sets of genes we found are RM systems that vary over the course of hours or days through a specific set of DNA rearrangements. This results in the patterns of methylation caused by the RM systems also changing over short timescales. Experimental data found that different forms of the inverting RM system caused different patterns of methylation, each of which was associated with a distinct pattern of gene expression. This epigenetic regulation of bacterial genes was found to change the virulence of the bacterium, with some patterns of methylation making the pneumococcus more likely to cause disease. This could be an important factor in the transition from the pneumococcus being a harmless commensal, to becoming a dangerous pathogen. This project is designed to test this hypothesis through studying whether the second variable RM system in the pneumococcus affects the same processes, or has a different effect on cell physiology, and whether such systems regulate the virulence of other pathogenic bacteria. Searching of the thousands of publically available bacterial DNA sequences has allowed us to identify hundreds of species that harbour similar systems. These include bacterial species that are very common in the human gut, some that are present in probiotic drinks and others involved in the production of cheese. Perhaps most importantly, they are also present in many pathogenic bacteria. This project is designed to investigate whether these variable RM systems might also regulate virulence in three bacterial species that each represent major threats to public health. The first is Streptococcus suis, a species normally associated with pigs that is emerging as a major pathogen capable of causing serious infections, such as meningitis. The second is Listeria monocytogenes, a foodborne bacterium that causes potentially fatal infections. The third is Enterococcus faecalis, a major cause of highly antibiotic-resistant infections, particularly in a hospital setting. In the three species, the variable RM loci are present with lineages that are associated with causing high levels of disease in humans, and absent from those that are asymptomatically found in animals or humans. The overall aim of the project is to work out how these systems may play a role in regulating genes involved in the bacteria's virulence, as well as how they evolved and how diverse they are. Such information will allow us to understand why these unusual genes are distributed, and why bacteria progress from being harmlessly carried to causing disease. This would better inform our strategies as to how to prevent this transition, and thereby tame these common, but potentially dangerous, bacteria.