Role of RNA repair in the tolerance of bacteria to antibiotics.

The killing of bacteria by antibiotics is preceded by a period of adaptation and tolerance to the antibiotic(s). Similarly sub lethal amounts of antibiotics cause adaptations in the bacterial cells allowing them to better survive in the presence of antibiotics. Identifying the mechanisms behind antibiotic tolerance will afford us with new ways to manage the administration of antibiotics for controlling infections, and also inform new strategies for enhancing the action of antibiotics. We have discovered certain antibiotics cause the induction of a repair system in bacterial cells. The system repairs particular molecules in the cell called RNA, which perform key functions for cell growth and maintenance. By working out how the cells use the RNA repair system to help them survive antibiotic challenges we will be able to plan how to use different specific combinations of antibiotics to kill the bacteria, and also embark on a program of work to consider inhibiting the repair system itself. We anticipate that we can potentiate the killing actions of existing antibiotics through careful establishment of appropriate combinatorial treatments of bacterial infections.

Understanding how antibiotics act on their target organisms is key to developing new knowledge based strategies to enhance and manage the action of existing antibiotics. Improving the efficiecies of existing antibiotics is one medium term route to reduce the emergence of resistance to antibiotics. We have shown certain antibiotics including Gentamicin (subject of MRC supported investigations to mitigate its effects on hearing in humans) cause the induction in E. coli bacteria of a conserved RNA repair system (the RtcAB proteins which are regulated by the AAA+ RtcR transcription activator protein), a physiological response to antibiotic challenges not previously recognized. Remarkably, some of the rtc-inducing antibiotics target the activity of ribosomes whereas others target the cells envelope synthesis and integrity. Initial experiments show the priming of the cells with the RNA repair system in its ON state changes the killing of cells by these antibiotics. We propose that the functional links between the translation apparatus and the cell envelope should be targeted for improved antimicrobial therapy. We aim to establish using RNAseq and antibiotic challenges in the presence and absence of the rtc system the identity of the key molecular players for the RNA repair mediated tolerance to antibiotics. Using temporal and spatial studies of the RtcR, A and B proteins we will arrive at the molecular signaling processes underlying activation of the repair system, which may then be targeted for inhibition by low molecular weight compounds (potentially including existing antimicrobials) and so allow us to potentiate the action of existing antibiotics.

We believe the major impacts will be on a range of beneficiaries The Key potential beneficiaries of the proposed research are:

(i) Academics: The academic sector will be the main short to medium term beneficiary, as the proposed research will provide knowledge, reagents and new imaging outcomes of a stress system linking the translation and transcription machinery in a major studied bacterium E. coli widely used to unravel the basic life processes for many decades as well as being a significant pathogen in its own right. Furthermore, the project will provide a clear opportunity for career development and training of individuals, both nationally and internationally.
(ii) Society at large: Benefits to society at large will be twofold: In the short term, the proposed project will provide employment and training for an individual at the advanced postdoctoral level providing experience of project design, management as well as its high level scientific implementation, thereby directly contributing to the national economy. The interdisciplinary nature of the proposed research will greatly enhance training of the associated PDRA, especially with respect to their ability to work within large interdisciplinary teams. Insights into the effective combinatorial use of antibiotics will benefit patients and health care professionals who treat them.
iii) Industry: The industrial sector is another potential long-term beneficiary. The proposed research will generate knowledge that could potentially be exploited for new product development by the pharmaceutical industry (e.g. against therapeutically proven anti-microbial targets, as anti-cancer agents where the Rtc system plays a role in humans) and stimulate development of appropriate delivery systems by the biomedical engineering sector, to include exploiting novel nucleic acid interacting activities in eg genome editing. Research results could potentially identify novel targets for therapeutic intervention at protein/RNA, protein/protein protein/DNA interaction level.The IC Business Development team would be a valuable resource in supporting any (long term) future commercial development arising from this research. Similarly, this would benefit from the expertise offered by IC Innovations in the area of translating research into marketable products.
iv) Government: One of the remits of the new IC Institute for Global Health is to translate new scientific knowledge into applications to improve global health by influencing international policy. Expertise offered by the IC Institute for Global Research could therefore be exploited for using discoveries made as a result of the proposed research to inform future health care policies.

Exploitation and Application: A number of structures exist within ICL for exploitation of knowledge gained and the development of beneficial applications. For example, we could make use of the expertise offered by ICL Innovations in the area of translating research into marketable products. In addition, we have the opportunity to benefit from input and advice from IC Drug Discovery Centre's multi-disciplinary team whose remit is to translate early research into drug discovery projects. Results from the project will provide opportunities for novel drug-target discovery centered around protein/RNA, protein/protein and protein/lipid interactions.

For drug discovery, and noting the diminishing content of the pipelines that feed this aim, one possibility is that, as a result of the proliferation of technologies intended to enable drug discovery, the basic biological questions are being overlooked or ignored. Technological development in high throughput target identification, screening, library synthesis, and validation have their place, but they are essentially just tools, and a clear understanding of the underlying biology is paramount.This project affords such an understanding