ATP-driven Spatial Organization of a Carbon-fixing Organelle in Cyanobacteria

Metabolic compartmentalization contributes to the spatiotemporal modulation of biochemical reactions within eukaryotic and prokaryotic cells. Bacterial microcompartments (BMCs) are widespread, self-assembling proteinaceous organelles that function by physically sequestering key metabolic enzymes essential in carbon source utilization, microbial ecology, and pathogenesis. A well-studied anabolic BMC, called the carboxysome, serves as a critical carbon-fixing organelle in cyanobacteria and many chemoautotrophs. Carboxysomes play a central role in the CO2-concentrating mechanism of these bacteria by compartmentalizing central photosynthetic enzymes, effectively increasing their efficiency, and preventing the occurrence of wasteful side reactions. The basic architecture of carboxysomes consists of multi-protein subunits that form a selectively permeable diffusion barrier, allowing gaseous and metabolic exchanges to take place. While eukaryotic organelles use cytoskeletal highways for their cellular organization, little is known about how bacteria spatially regulate their protein-based organelles for proper segregation, homeostasis, inheritance, and maintenance in a cell population. Fully assembled, mature carboxysomes are aligned equidistantly along the cell length to ensure faithful inheritance. Previous studies have implicated an important ParA-like protein we termed Maintenance of carboxysome distribution A (McdA) as a critical component in equal spacing of carboxysomes. McdA forms oscillating protein gradients over the nucleoid in response to its carboxysome-localized partner protein, McdB. Prior to my work, the mechanism driving McdA oscillations and carboxysome positioning by the McdAB system was unknown. To address this gap in knowledge I first examined the biochemical interaction between the two-component McdAB system and the nucleoid. I then dissected how the McdA ATPase cycle mediates the protein’s dynamic oscillatory patterning and carboxysome positioning. Finally, I demonstrate how multi-generational live-cell imaging of S. elongatus can serve as an indispensable tool in studying the mechanistic workings of carboxysome dynamics and inheritance. Together, this thesis work provides the first steps towards an understanding of the ATP-driven molecular mechanisms governing organelle trafficking in bacteria.PHDMolecular, Cellular, and Developmental BiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/171306/1/annhakim_1.pd