Characterization of Factors that Regulate Cell Envelope Synthesis in Staphylococcus aureus

Abstract

Bacteria are separated from their environment by a cell envelope, an essential structure composed of multiple polymers and proteins. To grow and divide, the existing structure needs to be remodeled then expanded with new material. These processes are inextricably linked: the degradation and synthesis of the cell envelope must be balanced to maintain its integrity. To accomplish this, cells have evolved a series of regulators in the cell envelope which can (i) directly modify the activities of cell envelope synthesis proteins or (ii) signal transduction proteins that can direct intracellular processes to respond to extracellular events. This thesis is split across two projects, each elucidating a regulatory complex in the membrane of S. aureus. In the first project, I describe a physical and genetic interaction between SpsB and SpbR (backgrounds for each protein are provided in Chapter 1.3). This project was undertaken with two other graduate students, Madeleine Stone and Youngseon Park, and together we describe how SpbR controls processing of LtaS to terminate lipoteichoic acid synthesis and maintain appropriate length LTAs in the membrane. In the second project, I describe a physical and genetic interaction between AuxB, PknB, and GpsB (backgrounds for each protein are provided in Chapter 1.4, Chapter 1.6, and Chapter 1.7). I show that AuxB forms independent interactions with GpsB and PknB; by altering the interaction between AuxB and PknB, I induce phenotypes that indicate the former regulates the latter. I also describe how the features of PknB and AuxB confer intrinsic resistance to various antibiotic compounds and have unique genetic relationships to cell envelope polymers. Finally, I present models for the possible physiological roles of AuxB in complex with PknB and GpsB. In Chapter 2, I characterize SpbR as an interacting partner of the signal peptidase SpsB and show that it regulates lipoteichoic acid synthesis by controlling cleavage of lipoteichoic acid synthase. In Chapter 3, I report and characterize AuxB-GpsB and AuxB-PknB interactions and present evidence that AuxB antagonizes PknB through its direct association. I further show that S. aureus becomes sensitive to various disruptions in cell envelope synthesis when the AuxBPknB complex is too abundant or lacking from the cell. In Chapter 4, I discuss unpublished work on AuxB and PknB that provides exciting hypothesis for future work. In Chapter 5, I discuss future directions and implications for this work