THE GENETICS OF METAL UPTAKE IN CAULOBACTER CRESCENTUS: CHARACTERIZATION OF REGULATORS AND IDENTIFICATION OF RECEPTORS

Caulobacter crescentus is an oligotrophic, Gram negative bacterium with a dimorphic lifecycle. It is a model system for cell cycle regulation and cellular development. Since C. crescentus resides primarily in freshwater, this microorganism can also serve as a model system for nutrient uptake by oligotrophic freshwater species. This study focused on TonB-dependent receptors and metal-dependent regulators in C. crescentus. To maintain appropriate levels of iron within the cell, bacteria have evolved mechanisms to control iron uptake, storage, and use. In microbes, there are two protein families possessing members that accomplish this: Fur and Rrf2. In the C. crescentus genome, there are two putative members from each family. I conducted a phylogenetic analysis to better understand the role of these four proteins. The Fur family proteins were most closely related to regulatory proteins that sense manganese (Mur-like) and zinc (Zur-like). In the Rrf2 family, one protein shared homology with IscR, which regulates Fe-S synthesis and utilization pathways, while the other protein apparently represents a new member of Rrf2 family of regulatory proteins. I found that this protein is produced in response to metal limitation. Future work on this complex topic will focus on determining which metal or metals each of these regulatory proteins responds to, as well as determining the binding sites. TonB-dependent receptors are responsible for transporting ferric-siderophore complexes into the periplasm of the cell. I was able to reduce the potential number of TonB-dependent receptors that might possess iron regulation to nine. However, results from another laboratory likely expand that number to thirteen. I found that C. crescentus is capable of utilizing at least five siderophores. Deletion analysis of three receptors, CC0028, CC1666, and CC3336, did not allow for linking a specific complex to a receptor. However, I found that CC0028 and CC3336 transcripts were up regulated 2.5 and 2.0 fold respectively in iron limited conditions. Interestingly, CC3336 is apparently regulated by both iron and carbon starvation. With one possible exception, this is the only TonB-dependent receptor seen to be regulated by two different stressors, one of which has been shown to utilize a sRNA