Regulation of elongation by RNA polymerase and ribosome via intrinsic signals and transcription-translation coupling

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Understanding the molecular mechanisms governing the expression of genes in the cell is an essential part of developing strategies for intelligent manipulation of harmful and useful organisms, development of new antimicrobial and antifungal agents, and, importantly, for understanding emergence and evolution of Life.In bacteria, both steps of gene expression, copying genes into RNA (transcription) and production of proteins on RNA (translation), are coupled. This implies tight interactions and mutual regulation of these two molecular machineries, which determine the overall rate of gene expression, which, in turn, is important for all cellular processes. This regulation and mechanisms determining the rates of transcription and translation are poorly understood. We are the first laboratory who developed a unique experimental system where both steps of gene expression can be assembled from purified components and investigated simultaneously in the test tube. We also possess some unique biochemical techniques, which allowed us for example to describe the mechanism involved in regulation of the development of HIV-1, and to uncover mode of action of antibiotic Tagetitoxin. In this proposal, by using these techniques, we will investigate what factors determine rates of both steps of gene expression: (i) we will apply a systematic approach to understand the signals in the DNA that may influence the rate of transcription; (ii) we will investigate the interactions and mutual regulation of transcription and translation. Sophisticated regulation of transcription and translation are the major factors for survival of pathogenic bacteria. Understanding of relations between these two steps of gene expression will improve our knowledge of these bacteria, and thus provide new tools for fighting infectious diseases. We collaborate with a bio-tech company on applying experimental tools developed in our research to elucidation of modes of action of new antibiotics, inhibitors of RNA polymerase. Being at the interface of transcription and translation, the proposed study will be valuable for both of these vast fields of molecular biology. We collaborate with the leading UK laboratories who will use the tools designed in this study to investigate mechanisms of gene expression by biophysical methods, such as single-molecule techniques, crystallography, cryo-electron microscopy. Furthermore, simultaneous assembly of the two major cellular machineries in the test tube may become the first step for production of artificially assembled living organisms.