The T6SS as a search engine for naturally validated antibacterial targets

T6SS 作为自然验证抗菌目标的搜索引擎

• 批准号: MR/S02316X/1
• 负责人: Alain Filloux
• 金额: $ 67.78万
• 依托单位: Imperial College London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2019
• 资助国家: 英国
• 起止时间: 2019 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FS02316X%2F1
• 关键词: T6SS; search; engine; naturally; validated

Bacterial resistance has emerged to all clinically relevant antibiotics. Despite the widespread recognition of this as a global crisis the majority of the strategies implemented thus far have been disappointingly unsuccessful, notably those based on rational drug design and target identification approaches. One fundamental and key issue is indeed to define what a good antimicrobial target is so that it would effectively be amenable to drug design. Over billions of years microbes have evolved the best ways to out-compete others to gain access to space and nutrients. They have identified bacterial Achille's heels and designed strategies to inactivate corresponding molecular pathways that would right away challenge bacterial growth and survival. This is what one can consider as naturally validated drug targets.The bacterial Type VI Secretion System (T6SS) is the ultimate weapon killing competitors by toxins injection. Most characterized T6SS toxins work against cell wall, membrane, nucleic acids, which are all classic targets for most of our current antibiotics. We are at the beginning of understanding how this system works and the characterized T6SS toxins represent only the tip of the iceberg. It is also important to realise that the bacteria that produces T6SS toxins would also produce specific immunity proteins that specifically protect against the activity of each individual T6SS toxin. One key observation is that the gene encoding the toxin and the gene encoding the immunity are found on the bacterial genome as tandem gene pairs next to each other.This project aims at exploiting the largely overlooked reservoir of natural antibacterials, namely the plethora of T6SS toxins that bacteria have evolved to effectively kill bacterial competitors and which we believe exist in a much larger number and have an unexpected broad range of diverse biochemical activity. While T6SS toxins themselves are unlikely to be effective therapeutics in the short term, the naturally validated antibacterial targets they point us towards would be of great value.A potent gram-negative bacterium using the T6SS to outcompete and kill foes is Pseudomonas aeruginosa, an organism which is high on the WHO list of pathogens that are critical for Anti-Microbial Resistance (AMR). Here, we will use P. aeruginosa as model to perform a systematic search for T6SS toxins, which are not identifiable using standard genomic and bioinformatic analysis. In previous work my laboratory implemented a genetic screen (TraDIS) to identify transposon mutants sensitive to the T6SS. These mutants are affected in distinct immunity genes and the characterization of the adjacent gene would lead to the identification of a novel T6SS toxin. Our screen was fully validated since it allowed to identify the toxin/immunity pairs which were already known and proved to be very effective at the identification of unsuspected T6SS toxins and their immunities. One of the T6SS toxin which we propose to fully characterize in the present proposal is called Tse8 and is suspected to interfere with protein synthesis, notably by hampering function of the transamidosome complex which leads to a shortage in Asn- and Gln-tRNA.Finding new toxins and determining their mechanisms of action is going to offer a gold mine of usable antibacterial targets that pharmaceutical companies would be able to consider in the future. Furthermore, understanding how bacteria fight each other in ways we might not suspect is what is needed to prepare translational impact.

细菌已经对所有临床相关的抗生素产生了耐药性。尽管人们普遍认识到这是一种全球危机，但迄今执行的大多数战略都令人失望地不成功，特别是那些基于合理药物设计和目标确定办法的战略。一个根本和关键的问题确实是定义什么是好的抗菌素靶标，以便有效地服从药物设计。在几十亿年的时间里，微生物进化出了最好的方式来击败其他微生物，获得太空和营养。他们已经确定了细菌的脚后跟，并设计了策略来灭活相应的分子途径，这些途径将立即挑战细菌的生长和生存。这就是自然有效的药物靶点。细菌VI型分泌系统(T6SS)是通过注射毒素杀死竞争对手的终极武器。最具特点的T6SS毒素对细胞壁、膜和核酸起作用，这些都是我们目前大多数抗生素的经典靶标。我们正在开始了解这个系统是如何工作的，T6SS毒素的特征只代表了冰山一角。同样重要的是要认识到，产生T6SS毒素的细菌也会产生特定的免疫蛋白，这种蛋白专门防止每一种T6SS毒素的活性。一个关键的观察结果是，编码毒素的基因和编码免疫的基因在细菌基因组上以串联基因对的形式被发现。该项目旨在开发被广泛忽视的天然抗菌素的储存库，即细菌进化出的过量的T6SS毒素，我们相信这些毒素存在的数量要大得多，并具有出人意料的广泛的不同生化活性。虽然T6SS毒素本身不太可能在短期内成为有效的治疗方法，但它们向我们指明的自然有效的抗菌靶点将是非常有价值的。一种使用T6SS来击败和杀死敌人的革兰氏阴性细菌是铜绿假单胞菌，这是一种在世卫组织对抗菌素耐药性(AMR)至关重要的病原体名单上排名靠前的微生物。在这里，我们将使用铜绿假单胞菌作为模型来进行系统的T6SS毒素的搜索，这些毒素无法用标准的基因组和生物信息学分析来识别。在以前的工作中，我的实验室实现了一个基因筛查(TRADIS)来识别对T6SS敏感的转座子突变体。这些突变体受不同免疫基因的影响，相邻基因的特征将导致一种新的T6SS毒素的鉴定。我们的筛查是完全有效的，因为它允许识别已知的毒素/免疫对，并被证明在识别未知的T6SS毒素及其免疫性方面非常有效。我们建议在本提案中完全描述的T6SS毒素之一称为Tse8，被怀疑可以干扰蛋白质的合成，特别是通过阻碍转氨体复合体的功能，从而导致ASN-和Gln-tRNA的短缺。发现新的毒素并确定其作用机制将为制药公司未来能够考虑的可用的抗菌靶点提供一个金矿。此外，了解细菌如何以我们可能不会怀疑的方式相互战斗是准备翻译影响所必需的。

项目成果

The Breadth and Molecular Basis of Hcp-Driven Type VI Secretion System Effector Delivery.

• DOI: 10.1128/mbio.00262-21
• 发表时间: 2021-06-29
• 期刊: mBio
• 影响因子: 6.4
• 作者: Howard SA;Furniss RCD;Bonini D;Amin H;Paracuellos P;Zlotkin D;Costa TRD;Levy A;Mavridou DAI;Filloux A
• 通讯作者: Filloux A

RpoN/Sfa2-dependent activation of the Pseudomonas aeruginosa H2-T6SS and its cognate arsenal of antibacterial toxins.

• DOI: 10.1093/nar/gkab1254
• 发表时间: 2022-01-11
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: Allsopp LP;Collins ACZ;Hawkins E;Wood TE;Filloux A
• 通讯作者: Filloux A

A novel stabilization mechanism for the type VI secretion system sheath.

• DOI: 10.1073/pnas.2008500118
• 发表时间: 2021-02-16
• 期刊: Proceedings of the National Academy of Sciences of the United States of America
• 影响因子: 11.1
• 作者: Bernal P;Furniss RCD;Fecht S;Leung RCY;Spiga L;Mavridou DAI;Filloux A
• 通讯作者: Filloux A

Causalities of war: The connection between type VI secretion system and microbiota.

• DOI: 10.1111/cmi.13153
• 发表时间: 2020-03
• 期刊: Cellular microbiology
• 影响因子: 3.4
• 作者: Allsopp LP;Bernal P;Nolan LM;Filloux A
• 通讯作者: Filloux A

Effectiveness of Pseudomonas aeruginosa type VI secretion system relies on toxin potency and type IV pili-dependent interaction.

• DOI: 10.1371/journal.ppat.1011428
• 发表时间: 2023-05
• 期刊: PLoS pathogens
• 影响因子: 6.7