Research
Sub-cellular Organization
Macromolecular Structures At Cell Poles
Every living thing has a body plan in which distinct anatomical parts are used for different purposes. This is true down to the fundamental unit of life - the cellular level. Even bacteria, the simplest and earliest known forms of cellular life, have defined body plans. Amazingly, bacteria can generate complex body plans within a single contiguous cytoplasm, without using internal membranes to provide compartmentalization or directed vesicular transport as eukaryotic cells do. Instead, bacteria can become organized through the assembly of complex macromolecular structures, which act as microdomains for the purpose of carrying out specialized functions. We seek to understand the organizational principles of a microdomains in the bacterium Caulobacter crescentus, which houses the control mechanisms for the establishment of cell polarity and the timing of chromosome replication in large macromolecular complexes at the cell poles. Using powerful genetic, biochemical, and visual tools afforded by bacterial systems, my laboratory will gain an atomic scale understanding of these microenvironments, and understand how their structure relates to biochemical mechanism. How is this dynamic arrangement of scaffolding proteins, signaling factors, and other molecules combined to form a functional macromolecular complex?
Cell Asymmetry and Bacteria as Multi-cellular Organisms
Asymmetric cell division entails the unequal separation of components between daughter cells. When this includes factors that regulate gene expression, the two daughter cells can run different genetic programs. Cellular asymmetry underlies basic developmental mechanisms such as tissue differentiation and stem cell maintenance, and is therefore critical for multicellular life. Interestingly, many bacteria are also capable of asymmetric cell division, and just as in animal cells, cell division produces two morphologically and transcriptionally distinct cell types - a basic form of multicellularity.
In Caulobacter crescentus, asymmetry is created by the establishment of two distinct multiprotein complexes at opposite cell poles. The localized proteins include at least seven histidine kinases and response regulators, a protease complex, and a host of other factors, and all of these work together to establish a robust pattern of asymmetric gene expression in the daughter cells. Such a sophisticated mechanism must have originated from a simpler form, but its complexity makes it difficult to deduce the basic framework. What are the minimal components needed to establish cellular asymmetry and generate differential gene expression, and how could such a mechanism have evolved?
An attractive starting point for addressing these questions is the polar organizing protein PopZ. PopZ is an essential component of the polar multiprotein complexes in Caulobacter crescentus. The gene is conserved through alpha-proteobacteria, indicating inheritance from an early stage of prokaryotic evolution.
Interestingly, PopZ shows a strong tendency to accumulate at a single pole when expressed heterologously in E. coli, an unrelated species without robust asymmetry or cell polarity. We are testing the hypothesis that PopZ was an early player in the evolution of cellular asymmetry in the alpha-proteobacterial lineage, and that it is part of a core polarity mechanism from which more elaborate systems have evolved. Can PopZ be used to build simple polarized signaling complexes in E. coli and can they be used to drive asymmetric gene expression in this system?
Synthetic Biology
Caulobacter crescentus cell poles are microenvironments in which protein interactions and other chemical reactions are reactions are concentrated and controlled in a space that is separate from the surrounding cytoplasm. Using our growing knowledge of the PopZ polar assembly scaffolding protein, its binding partners, and the organization of proteins within this complex network, will it be possible to re-engineer this microenvironment to perform novel functions?
