Signal transduction pathways allow organisms to sense and respond appropriately to a complex bouquet of environmental cues. The molecular determinants of specificity are constrained by the demands of signaling fidelity, yet flexible enough to allow pathway remodeling to meet novel environmental challenges. A detailed picture of how these forces shape bacterial two-component signaling systems has emerged over the last decade. However, the tension between constraint and flexibility in more complex architectures has not been well-studied. In this thesis, I combine comparative genomics and in vitro phosphotransfer experiments to investigate pathway remodeling using the Firmicutes sporulation initiation (Spo0) pathway as a model. The present-day Spo0 pathways in Bacilli and Clostridia share common ancestry, but possess different architectures. In Clostridia, a sensor kinase phosphorylates Spo0A, the master regulator of the sporulation, directly. In Bacilli, Spo0 is phosphorylated/activated indirectly via a four-protein phosphorelay. The presence in sister lineages of signaling pathways that activate the same response regulator and control analogous phenotypes, yet possess with different architectures, suggests a common ancestral pathway that evolved through interaction remodeling. The prevailing theory is that the ancestral pathway was a simpler, direct phosphorylation architecture; the more complex phosphorelay emerged within the Bacillar lineage. In contrast to this prevailing view, my analysis of 84 representative genomes supports a novel hypothesis for the evolution of Spo0 architectures, wherein the two protein, direct phosphorylation architecture is a derived state, which arose from an ancestral Spo0 phosphorelay. The combination of my bioinformatic analysis and the first experimental characterization of a Clostridial phosphorelay provide evidence for the presence of functional phosphorelays in both classes Bacilli and Clostridia. Further, a cross-species complementation assay between phosphorelays from each class suggests that interaction specificity has been conserved since the divergence of this phylum, 2.7 BYA. My results reveal a patchy phylogenetic distribution of both Spo0 pathway architectures, consistent with repeated remodeling events, in which a phosphorelay was replaced with a two protein, direct phosphorylation pathway. This remodeling likely occurred via acquisition of a sensor kinase with direct specificity for Spo0A. Further, my analysis suggests that the unusual architectures of the Spo0 pathway and its striking tendency to gain and lose interactions may be due to the juxtaposition of three key properties: the maintenance of interaction specificity through molecular recognition; the ecological role of endosporulation; and the degeneracy of interaction space that permits the ongoing recruitment of kinases to recognize novel environmental signals.
| Identifer | oai:union.ndltd.org:cmu.edu/oai:repository.cmu.edu:dissertations-2065 |
| Date | 01 August 2017 |
| Creators | Davidson, Philip |
| Publisher | Research Showcase @ CMU |
| Source Sets | Carnegie Mellon University |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Dissertations |
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