Understanding Spatial and Temporal Organization of Cyanobacteria for Synthetic Biology Applications

Abstract

The goal of synthetic biology is to engineer biological systems in order to solve industrial and medical challenges, as well as to learn about these systems by building. Cyanobacteria, a chassis for such engineering, are major players in the global carbon cycle and their ability to fix carbon has been harnessed to produce various chemicals, including biofuels. In addition, cyanobacteria possess remarkable spatial and temporal organization in the cell. In this dissertation, I monitor, break down, and rebuild the molecular components necessary for this spatial and temporal coordination of cyanobacterial growth. These studies give us a better understanding of basic cyanobacterial biology and enable the further development of cyanobacteria for synthetic biology applications. In chapter 1, I describe the arrangement of the cyanobacterial chromosomes over time, showing the mechanisms regulating chromosome duplication and segregation. The polyploid nature of cyanobacteria make this study relevant to their efficient genome engineering. In chapter 2, I elucidate the assembly of the primary carbon fixation machinery, carboxysomes. I show that the internal cargo of carboxysomes, RuBisCO, seeds assembly, followed by the recruitment of shell proteins, which form a solvent-protected microenvironment. Finally, in chapter 3, I engineer a synthetic circadian clock from cyanobacterial components in a heterologous organism, E. coli. I demonstrate the clock's modularity and pave the way for its use in medical and industrial applications. Taken together, this work furthers our understanding of cyanobacterial physiology and forms a foundation for their efficient engineering to increase their carbon fixation capabilities. Furthermore, the fundamental spatial and temporal organization strategies elucidated here can serve as inspiration for the engineering of heterologous systems that can serve similar purposes in different contexts.