Coordinated Spatial Organization in Bacteria

Subcellular organization in eukaryotic cells is largely governed by linear motor proteins. In bacteria, where linear motors are absent, a widespread family of ParA/MinD (A/D) ATPases are responsible for spatially organizing an array of genetic- and protein-based cellular cargos, the majority of which are unstudied. Of the well-studied examples, ParA ATPases segregate chromosomes and MinD ATPases position cell division in bacteria. Less studied is the growing list of A/D ATPases that spatially organize diverse protein-based cellular cargos, such as bacterial microcompartments (BMCs), flagella, chemotaxis arrays, and conjugation machinery. The long term goal is to understand how A/D ATPases drive and coordinate the positioning of such a diverse set of fundamental cellular cargos across bacteria, including those important to human health. Preliminary data from the project team shows that over a third of all bacteria encode multiple A/D ATPases. Among these bacteria are several human pathogens. The project team identified a nonpathogenic and experimentally tractable bacterium, Halothiobacillus neapolitanus, encoding six A/D ATPases, where each ATPase is dedicated to the positioning of a specific cellular cargo: the chromosome, the divisome, the carboxysome BMC, the flagellum, the chemotaxis array, and the conjugation machinery. The goal for the next five years is to use H. neapolitanus as an ideal model for probing mechanistic commonality, variation, and coordination, among the most widespread ATPase family used in the subcellular organization of bacterial cells. These advancements are important for human health because several of the bacteria encoding multiple A/D ATPases have associated cargos involved in pathogenesis. The overall objective here is to determine how multiple A/D ATPases coordinately function to spatially organize diverse cellular cargos. The central hypothesis is that in bacteria encoding multiple A/D ATPases, each ATPase is dedicated to the positioning of a specific cellular cargo, with specificity provided by an adaptor protein that links the cargo to its cognate ATPase, and that these positioning reactions are spatiotemporally coordinated. The rationale for the proposed research is that by determining how multiple A/D ATPases coordinately function within a single bacterial organism the project team will define the general mode of transport among this ATPase family, determine how the mechanism is altered for disparate cargos, and identify the specificity determinants for each cargo. The proposed research is creative and original because it has only recently been appreciated that protein-based organelles are prevalent across the bacterial world, making organelle trafficking in bacteria a new field of study. The project team will leverage this knowledge in the design of minimal and modular positioning systems for the faithful inheritance of cellular cargos in heterologous bacteria for synthetic biology approaches and biomedical application. Finally, the project team anticipates these findings will inform identification of novel positioning mechanisms and antibiotic targets important for bacterial pathogens.