Regulation of energy metabolism in two sulfur oxidizing Chlorobiaceae isolates
Date
2018
Authors
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Publisher
University of Delaware
Abstract
The green sulfur bacteria (Chlorobiaceae) are anaerobes that use electrons from reduced sulfur compounds (sulfide, S(0), and thiosulfate) as electron donors for photoautotrophic growth. Little is known in terms of the mechanics of gene expression regulation in energy metabolism or other cellular processes in the Chlorobiaceae. The work in this thesis expands our knowledge of how the Chlorobiaceae mechanistically regulate their gene expression, and thereby their physiology, in respect to energy metabolism. ☐ Chlorobaculum tepidum, the model system for the Chlorobiaceae, both produces and consumes extracellular S(0) globules depending on the availability of sulfide in the environment. These physiological changes imply significant changes in gene regulation, which has been observed when sulfide is added to Cba. tepidum growing on thiosulfate. A putative polysulfide oxidoreductase complex, psrABC, was shown to be highly upregulated on sulfide relative to thiosulfate and S(0). In this work, the first markerless mutagenesis system was developed and implemented in Cba. tepidum to delete the psrABC operon. Physiological characterization of the ∆psrABC mutant suggests that this protein complex confers high sulfide tolerance. ☐ While significant changes in gene expression were observed for Cba. tepidum in response to sulfide, a mechanistic understanding of how Cba. tepidum regulates its gene expression was lacking. Here, differential RNA-seq (dRNA-seq) was used to globally identify transcript start sites (TSS) that were present during growth on sulfide, biogenic S(0), and thiosulfate as sole electron donors. TSS positions were used in combination with RNA-seq data from cultures growing on these same electron donors to identify both basal promoter elements and motifs associated with electron donor dependent transcriptional regulation. These motifs were conserved across homologous Chlorobiaceae promoters. Together, this data suggest that sulfide-dependent repression is the dominant mode of gene regulation in Cba. tepidum, and wider Chlorobiaceae, and significantly enhances our understanding of the mechanisms of gene expression regulation in these bacteria. ☐ The Chesapeake Bay undergoes seasonal anoxia in the upper and middle portions, and when physical mixing occurs, sulfide fluxes enter the water column, creating anoxic, sulfidic waters. The Chlorobiaceae have been implicated in being the agents responsible for the oxidation of this sulfide. A Chlorobiaceae-dominated enrichment from a field site in the Chesapeake Bay was shown to be made up of Prosthecochloris sp. CB11, a green sulfur bacteria capable of sulfide oxidation rates at very low light levels. The genome and transcriptome of Prosthecochloris sp. CB11 grown under high and low light fluxes were sequenced. Many of the differentially expressed genes were related to growth rate control, biosynthesis, and S(0) metabolism. Two prophages, covering 40 open reading frames (ORFs) and 90 ORFs, were discovered in the genome, and represent the most complete Chlorobiaceae prophages to date. This data increases our understanding of how the Chlorobiaceae regulate their physiology in response to different energy regimes, and gives insight into how CB11 functions in the Chesapeake Bay. ☐ Together, this work increases our understanding of the regulation of energy metabolism in the Chlorobiaceae. In addition, a markerless mutagenesis system was developed that will aid in future studies for the characterization of the genetic components identified in this and future studies.
Description
Keywords
Biological sciences, Environmental microbiology, Green sulfur bacteria, Metabolism, Microbial physiology, Regulation, Systems biology