Understanding and manipulating a conserved and essential transcription factor to activate antibiotic production in Streptomyces species

人类医学中使用的几乎所有抗生素都是在50年前发现的，大多数致病细菌现在对其中一种或多种药物具有抗药性。这意味着人类正面临着回到抗生素出现前的时代，这一令人担忧的情况被描述为“抗生素末日”。政府委托的奥尼尔抗菌素耐药性审查(AMR)估计，如果我们现在不启动抗生素发现工作，那么到2050年，耐药感染导致的死亡人数将超过癌症，估计每年有1000万人死亡。我们目前使用的大多数抗生素都是从土壤细菌的次生代谢物中提取的天然产物，最重要的一类是链霉菌，它制造了所有已知抗生素的50%。链霉菌对人类非常重要，尽管科学家已经从这些细菌中发现了许多抗生素，但我们现在知道，他们只发现了真正容易找到的化合物--低垂的果实。过去15年的基因组测序显示，链霉菌在实验室条件下只产生约25%的次生代谢物，这意味着从1940年到1960年，也就是所谓的抗生素发现的黄金时代，科学家们几乎没有对它们的能力进行采样。其余的被称为“沉默的”次生代谢物，因为它们不是在实验室中产生的。好消息是，与20世纪的科学家相比，这意味着我们有很大的优势--如果我们能找到方法来生产已知的600个物种中所有沉默的次生代谢物，我们就会找到许多新的抗生素，可以进入临床试验的管道。这是抗生素发现的最早阶段，我们现在加大努力是至关重要的，因为需要10-15年的时间才能使药物通过临床试验并获准在人体上使用。可能有1%的抗生素适合治疗疾病，所以在接下来的几十年里，我们从链霉菌中发现的天然产品越多越好。激活沉默的次生代谢物产生的一种方法是了解控制其在土壤中产生的自然信号和信号通路，这是我们研究的重点。如果我们能够操纵这些信号通路，我们就可以迫使细菌在实验室里制造所有的抗生素。理想情况下，我们希望确定在所有600多种已知链霉菌中影响抗生素生产的信号通路，这是我们建议的主题。我们已经确定了一条由两种蛋白质组成的信号通路，称为MTRA和MtrB，并发现这是链霉菌属中唯一保守和必要的通路。这意味着这种MtrAB双组分系统在每一株测序的链霉菌中都能找到！MTRA是一种DNA结合蛋白，其活性受信号感应蛋白MtrB控制。如果我们通过删除MTRA基因来扰乱这一途径，那是致命的。如果我们删除mtrB基因，它就不需要环境信号来激活该途径，并导致活性MTRA蛋白的过度生产，从而启动抗生素的生产，而抗生素在野生型菌株中通常是沉默的。然而，单纯的过度生产MTRA是行不通的，我们也必须取消MTRA。在这个项目中，我们将分析两个模式物种的MtrAB，它们分别是天蓝色链球菌和条斑链球菌。我们将确定MtrB如何控制MTRA的活性，为什么MTRA在没有MtrB的情况下是活跃的，以及MTRA为什么以及如何激活沉默的次生代谢物的产生。我们还将试图获得总是活跃的功能MTRA蛋白，并看看我们是否可以用它们来开启我们的模式菌株和我们分离并测序的两个新的有才华的链霉菌物种的抗生素生产。我们称它们为天才，因为它们似乎编码了许多新的次生代谢物，而MTRA可能会让我们从这些菌株中发现新的抗生素。

Almost all the antibiotics used in human medicine were discovered >50 years ago and most disease-causing bacteria are now resistant to one or more of these drugs. This means humans are facing a return to the pre-antibiotic era, an alarming situation that has been described as 'antibiotic Armageddon'. The government commissioned O'Neill review on AntiMicrobial Resistance (AMR) estimates that if we do not kick-start antibiotic discovery efforts now then by 2050 drug resistant infections will kill more people than cancer, an estimated 10 million a year. Most of the antibiotics we currently use are natural products derived from the secondary metabolites of soil bacteria and the most important group are called Streptomyces, which make 50% of all known antibiotics. Streptomyces are incredibly important to humans and although scientists have already discovered lots of antibiotics from these bacteria we now know that they only found the really easy to find compounds, the low hanging fruit. Genome sequencing over the last 15 years has revealed that Streptomyces bacteria only make about 25% of their secondary metabolites under laboratory conditions which means that from 1940-60, the so-called golden age of antibiotic discovery, scientists were barely sampling their capability. The rest are called 'silent' secondary metabolites because they do not make them in the lab. The good news is this means we have a big advantage over scientists working in the 20th century - if we can find ways to switch on production of all the silent secondary metabolites in the >600 known species we will find lots of new antibiotics that can enter the clinical trials pipeline. This is the earliest stage in antibiotic discovery and it is vital that we increase our efforts now because it takes 10-15 years to get drugs through clinical trials and approved for use in humans. Probably <1% of antibiotics will be suitable for treating disease so the more natural products we can discover from Streptomyces in the next few decades the better. One way to activate the production of silent secondary metabolites is to understand the natural signals and signalling pathways that control their production in the soil and this is the focus of our research. If we can manipulate those signalling pathways we can force the bacteria to make all of their antibiotics in the laboratory. Ideally we want to identify signalling pathways which effect antibiotic production in all 600+ known Streptomyces species and this is the subject of our proposal. We have identified a signalling pathway consisting of two proteins called MtrA and MtrB and found this is the only conserved and essential pathway in the genus Streptomyces. This means this MtrAB two-component system is found in every single sequenced Streptomyces strain! MtrA is a DNA binding protein and its activity is controlled by the signal sensing protein MtrB. If we disrupt the pathway by deleting the mtrA gene it is lethal. If we delete the mtrB gene it removes the need for an environmental signal to activate the pathway and results in over-production of active MtrA protein which switches on production of antibiotics that are usually silent in the wild-type strains. However, simply over-producing MtrA does not work, we HAVE to remove MtrB as well. In this project we will analyse MtrAB in two model species called S. coelicolor and S. venezuelae. We will determine how MtrB controls MtrA activity, why MtrA is active in the absence of MtrB and why and how MtrA activates the production of silent secondary metabolites. We will also try to make gain of function MtrA proteins that are always active and see if we can use them to switch on antibiotic production in our model strains and in two new talented Streptomyces species that we have isolated and genome sequenced. We call them talented because they appear to encode many novel secondary metabolites and MtrA may allow us to discover new antibiotics from these strains.