Unraveling how N. meningitidis distinguish self from non-self DNA during DNA transformation

自然转化是指细菌（定义为遗传活性细菌）从细胞外环境中摄取裸DNA并通过重组将其纳入其基因组的过程。至少有60种细菌是自然转化的，这个数字被认为是相当低估的。自然转化是一种非常有价值的研究工具，有助于生物学中最重要的发现之一（即遗传物质是DNA）。此外，现在很清楚，通过自然转化的水平基因转移是产生遗传多样性的强大机制，也是细菌进化的主导力量。IV型皮利（Tfp）或相关的细胞器在DNA转化中起关键作用。此外，它们是细菌中最广泛的定植因子之一。因此，Tfp已经并将继续被深入研究，因为它们可能是开发针对细菌病原体的新疗法的主要靶点，这些病原体通过感染人类、牲畜和农作物给人类健康和经济带来沉重负担。在目前的模型中，裸DNA需要在菌毛缩回时被细菌吸收之前被Tfp结合，但这仍有待正式证明。Tfp中DNA受体的存在由于以下事实而得到加强：在某些物种中，只有自身DNA被摄取，因为它含有被称为摄取序列的特征序列基序。在奈瑟氏球菌中，这种DNA摄取序列（DUS）基序长10 bp，存在于约10个核苷酸中。每个基因组有2,000个拷贝有能力的细菌如何结合DNA并能够在摄取过程中区分自我和非自我DNA仍然是一个谜。DUS受体的鉴定和分析将代表一个重大突破，并将提高我们对DNA转化和Tfp生物学的理解。Tfp生物学相关的脑膜炎病毒蛋白。这些蛋白质中的三种（ComP、PilV和PilX）是Tfp的次要（低丰度）组分。竞争力在竞争力中起着关键作用。这促使我们纯化ComP蛋白并测试其DNA结合活性，这导致了一个重大发现：ComP具有DNA结合活性并显示出对DUS的偏好（未发表的数据）。在这个项目中，我们计划使用生物化学和细胞分析以及高分辨率结构技术来彻底表征ComP的DNA结合活性。

Natural transformation is the process by which bacteria, defined as genetically competent, take up naked DNA from the extracellular environment and incorporate it into their genomes by recombination. At least 60 bacterial species are naturally transformable, a number that is thought to be considerably underestimated. Natural transformation is an extremely valuable tool for research and was instrumental to one of the most important discoveries in biology (i.e. that the genetic material is DNA). Moreover, it is now clear that horizontal gene transfer by natural transformation is a powerful mechanism for generating genetic diversity and a dominant force in bacterial evolution. This can have devastating consequences, e.g. by promoting spread of virulence or antibiotic resistance genes.Type IV pili (Tfp), or related organelles, play a key role in DNA transformation. In addition, they are one of the most widespread colonization factors in bacteria. Therefore, Tfp have been and continue to be intensively studied as they might be primary targets for the development of new therapies against bacterial pathogens that place a heavy burden on human health and economy by infecting mankind, livestock and crops.In current models, naked DNA needs to be bound by Tfp before it is taken up by the bacteria upon pilus retraction, but this remains to be formally demonstrated. The existence of a DNA receptor in Tfp is strengthened by the fact that in some species only self DNA is taken up because it contains a signature sequence motif known as the Uptake Sequence. In Neisseria, this DNA Uptake Sequence (DUS) motif is 10 bp long and is found in approx. 2,000 copies per genome. How competent bacteria bind DNA and are able to discriminate self from non-self DNA during uptake remains a mystery. The identification and analysis of a DUS receptor would represent a major breakthrough and would improve both our understanding of DNA transformation and Tfp biology in general.This proposal originally derives from our recent characterization of N. meningitidis proteins involved in Tfp biology. Three of these proteins (ComP, PilV and PilX) are minor (low abundance) components of the Tfp. ComP plays a key role in competence. This prompted us to purify the ComP protein and test its DNA-binding activity, which led to a major finding: ComP has DNA-binding activity and shows a preference for the DUS (unpublished data). During this project, we plan to thoroughly characterize ComP's DNA-binding activity using biochemical and cellular assays, together with high resolution structural techniques.