Physical model of the bacteria nucleoid

Project Summary The bacterial genome is organized into a compact form, known as the nucleoid, consisting primarily of the supercoiled circular DNA chromosome and DNA-binding proteins. Generation and maintenance of the nucleoid structure, which arises without a nuclear membrane or the chromatin present in eukaryotes, is critical to bacterial health; understanding the factors that give rise to this structure are thus important for promoting synergistic bacteria and disrupting antagonistic pathogens. Likewise, the dynamics governing DNA loci diffusion and intranucleoid transport govern the time scales for DNA-protein binding. A key open question surrounding the structure and dynamics of the nucleoid is elucidating those factors that can be attributed to its polymeric nature and those that are of biological origin. Indeed, many of the most intriguing ideas emerging from the microbiology community in the past decade, such as density waves, the glass-like behavior of the nucleoid in the absence of metabolic activity, and the connection between solvent quality and macrodomain formation, rely on connecting nucleoid biology to polymer physics. Unfortunately, making these connections in a definitive, quantitative manner has been frustrated by (i) the lack of a suitable physical model of the nucleoid and (ii) experimental data that, when combined with such a model, are sufficient to establish the governing biophysical principles. To resolve this gap in our understanding, this proposal couples (i) a multiscale, polymer physics-based model of the nucleoid that reflects both its equilibrium and non-equilibrium aspects to (ii) state-of-the-art biophysical experiments probing the nucleoid structure and dynamics. Any model of the nucleoid must respect the biological fact that the nucleoid is not a fully equilibrated system; the combination of topological domains, macrodomains and chromosome interacting domains are evidence that the nucleoid is not in a global equilibrium that samples the full configurational space, even if these domains can locally equilibrate over the time scale of cell division. We will parameterize a computationally tractable, coarse-grained (CG) model of the entire out-of- equilibrium nucleoid based on simulations of the rapidly equilibrated, fine-scale phenomena. By construction, the CG model will incorporate large-scale, non-equilibrium constraints imposed by biology. We will then study nucleoid structure and dynamics over parameter ranges far beyond those available in existing data sets, using both GPU-accelerated simulations of the CG model, live cell imaging, and a microfluidic experimental system. The experiments will improve the CG model through feedback between them, while the CG model will reveal the most useful systems for experimentation and aid in interpreting their results. Our approach will definitively establish the physical laws governing nucleoid structure and dynamics. Applying those laws at biologically relevant parameter ranges then provides a foundation for identifying when polymer physics can (or, even more importantly, cannot) be used to understand nucleoid biology.