Spatial-temporal control over tipping-point operation defines fidelity of genome partition

Johns Hopkins University, Baltimore MD

Investigators

Abstract

Cells (especially bacteria) exist and thrive under a range of environmental conditions and they must alter various functions to adapt to changing conditions. This is challenging because many processes operate in a semi-stable state. This means that many processes can go one way or another, depending upon a tipping point (or operating point) which upon activation leads the cell down the proper path. Thus, cellular processes must establish the right operating point to robustly execute function and at the same time,¬ sensitively adapt this point to changing environmental cues. The overarching goal of this project is to elucidate how cells control the near-tipping-point operation to ensure the fidelity of cellular processes. This project will examine genetic partitioning in bacteria using a combined computational modeling and experimental approach to reveal how cells segregate DNA under changing conditions. The project will shed important light on how evolution shapes the operating point of this important cellular process to ensure genetic material is partitioned with high fidelity. The project will include training of undergraduate and graduate student researchers, community outreach efforts promoting the academic representation of underrepresented minorities, and creating a new course, “Mechanistic modeling of cell biology”, which emphasizes how to meaningfully integrate modeling with experiments. An annual workshop on the emerging topics will also be developed and will challenge participants to begin developing reasonable models of essential processes. Low-copy plasmid partitioning in bacteria provides a well-suited paradigm to distill the fundamental principles underlying the fidelity of genome partitioning. Most low-copy plasmids are actively segregated along the nucleoid by the conserved tripartite ParABS system. Upon replication, the sister plasmids always segregate by about half of the nucleoid length, ensuring high fidelity partitioning into the two daughter cells. As plasmid partitioning is not coupled to cell cycle and the nucleoid keeps elongating before cell division, the sister plasmids can be anywhere along the nucleoid when the parental cells starts to divide. This precipitates the key unanswered question of how the ParABS-mediated partition faithfully adapts the plasmid segregation distance to half of the nucleoid length to ensure plasmid partition fidelity. The project team established: 1) bacterial low-copy plasmid partitions via a Brownian ratchet mechanism where the plasmid “self-drives” by creating and following the ParA concentration gradient on nucleoid, and; 2) this ratcheting in vivo operates near a tipping point in the parameter space. The project will have two specific aims: 1) determine how ParA-mediated partition sensitively adapts to the length of the elongating nucleoid, and 2) establish how ParA-mediated partition ensures robustness of plasmid segregations. The project will elucidate how the ParA spatial-temporal regulation controls this near-tipping-point operation and ensures the fidelity of genome partition against stochastic fluctuations (e.g., the variations in ParA level). The basic principles distilled from this project would help address one fundamental question in cell biology: How do cells faithfully measure cellular-scale distance by using only molecular-scale interactions? This project is partially supported by the Genetic Mechanisms cluster in the Division of Molecular and Cellular Biosciences. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.