Date of Award
12-2023
Document Type
Campus Access Dissertation
Degree Name
Doctor of Philosophy (PhD)
Department
Biology/Molecular, Cellular, and Organismal Biology
First Advisor
Katherine E. Gibson
Second Advisor
Alexey Veraksa
Third Advisor
Linda Huang, Adan Colon-Carmona, Graham Walker
Abstract
The cell cycle is a fundamental process involved in bacterial reproduction and cellular differentiation. For Sinorhizobium meliloti, its cell cycle outcomes depend on its growth environment. This bacterium shows tight coupling of DNA replication initiation and cell division during free living growth, as well as asymmetric daughter cell fate. In contrast, it undergoes a program of endoreduplication and terminal differentiation during symbiosis with its host. While several key cell cycle components of the CtrA TCS pathway have been identified as playing an important role in this differentiation process, there is a lack of resolution regarding the molecular activities that are required and whether they could be unique to this cell type. CbrA, a negative regulator of CtrA activity, is absolutely required for a successful symbiosis. In this work, spontaneous symbiosis suppressors of the ∆cbrA mutant identified alleles of divL and cckA. In addition to rescuing symbiosis development, they rescue free-living cell cycle phenotypes of the ΔcbrA mutant. Moreover, our investigation into ∆cbrA polyploidy revealed this phenotype is most likely due to a delay in cell division rather than excessive DNA replication initiation events. Biochemical characterization of the hybrid histidine kinase CckA in vitro demonstrates that it has both kinase and phosphatase activities. In particular, CckA phosphatase activity is induced by the second messenger c-di-GMP. While the suppressor CckAA373S appears to maintain phosphatase activity in response to c-di-GMP in vitro, it has a significant loss in kinase activity that would result in decreased CtrA~P activity in vivo. The findings presented in this work deepen our understanding of the CtrA TCS pathway and open new avenues for further molecular characterization of a regulatory network pivotal to the free-living cell cycle and bacterial symbiosis.
Recommended Citation
Bender, Hayden A., "Molecular Mechanisms Governing Symbiosis and Cell Cycle Progression: Analysis of DIVL/CCKA Suppressor Alleles in Sinorhizobium Meliloti" (2023). Graduate Doctoral Dissertations. 898.
https://scholarworks.umb.edu/doctoral_dissertations/898
Comments
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