Characterization of novel envelope proteins and their relationship with the Cpx response and stress resistance in Escherichia coli
Creators: Bishop, Justin G
Publisher: name: University of Alberta Library
Year: 2022

<p>Stress is a major factor every organism needs to mitigate to survive. Environmental factors such as oxygen content, temperature, and alkalinity can all induce stress and prevent life from progressing. For Escherichia coli and other members of the mammalian gastrointestinal tract they need to survive fluctuations in pH, osmolarity, and nutrient availability all while fighting off the host immune system, bacteriophages, and other microbes. The bacterial envelope, the first layer of protection against external stress, is a physical barrier separating the external environment from the cell’s cytosol and acts as a matrix for respiration, transport, and the proton motive force. If the envelope fails, cellular processes fail. Envelope homeostasis is maintained by a suite of envelope stress responses that sense and respond to perturbations through the regulation of various envelope processes. E. coli possess five major envelope stress responses that work together to mitigate stress at the various membrane layers. The Cpx stress response is a crucial component that helps maintain envelope homeostasis at the inner membrane and periplasmic space through the regulation of inner and transmembrane complexes, chaperones, and proteases. Regulon analysis reveals the Cpx response influences a broad set of cellular processes ranging from virulence to metabolism. The Cpx response’s origins lie in the assembly of the conjugative pilus. Conjugation is one of three methods of horizontal gene transfer, the others being natural transformation and transduction by bacteriophages. Some bacteriophages can integrate into the host genome as prophages and have been shown to aid in resistance to various stress, influence host gene regulation, and provide a competitive advantage over other bacterial cells. In this study, our aim was to search for a link between prophage encoded genes in enteropathogenic E. coli strain E2348/69 and the Cpx response to determine if they contributed to mitigation of envelope stress. We found that E2348/69 contains active prophages with a narrow host range. We cross referenced the prophage regions in E2348/69 with a transcriptome<br>
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dataset collected during Cpx activation and located three putative genes (yfgG, yfgH, and yfgI) near, but not encoded by prophage region 10. Further examination of these genes revealed a putative CpxR binding site upstream of yfgG and gene expression analysis showed the expression of yfgG, and possibly yfgH, were influenced by the Cpx response. We found that these proteins are localized to the envelope and aid in resistance to alkaline, cobalt, and nutrient limitation in conjunction with the Cpx response. Absence of the outer membrane lipoprotein YfgH strongly induced the Cpx response in a CpxP dependent manner, a novel finding in the field. Transcript and protein analysis revealed an increase in cellular CpxP levels and periplasmic DegP levels. Together our data suggests these envelope localized proteins are key factors in envelope homeostasis mediated by the Cpx response.</p>