Regulation of Respiration by the Cpx Response in Enteropathogenic Escherichia coli

• Author / Creator

  Guest, Randi L

• Gram-negative bacteria are separated from the environment by a multilayered structure known as the envelope, which is comprised of the outer membrane, the inner membrane and the aqueous periplasmic space that lies between the two membranes. Biogenesis of the envelope is a complex process that requires the coordinated activity of several different processes. To maintain a contiguous barrier, Escherichia coli utilize several signal transduction pathways that sense and mediate adaptation to defects in envelope biogenesis or envelope integrity. One such pathway is the Cpx envelope stress response, which mediates adaptation to stresses that affect protein folding within the envelope. To alleviate protein-folding stress, the Cpx response activates the transcription of several envelope-localized chaperones and proteases while also repressing the expression of macromolecular envelope protein complexes. While it is clear that regulation of chaperones and proteases would generate an effective adaptive response to misfolded proteins, the role of multisubunit protein complexes in this process is not yet known. Recent analysis of the Cpx regulon in enteropathogenic E. coli suggests that the Cpx response inhibits the expression of several respiratory complexes of the aerobic electron transport chain. The purpose of this thesis was to better understand the role of respiration in the Cpx envelope stress response, with a focus on the primary dehydrogenase NADH dehydrogenase I and terminal oxidase cytochrome bo3. First, we provide evidence to suggest that the Cpx response regulates expression of these complexes at the transcriptional and post-transcriptional level. Transcription of the genes encoding both complexes is directly repressed by the Cpx response. Furthermore, we found that activation of the Cpx response increases proteolysis of both NADH dehydrogenase I and cytochrome bo3 in a manner that may involve the zinc metalloprotease FtsH. We also show that regulation of these complexes facilitates adaptation to several stresses that activate the Cpx response. Next, we investigated the relationship of respiration to the stress that is sensed by the Cpx response. We found that both NADH dehydrogenase I and cytochrome bo3 contribute to basal levels of Cpx pathway activity, but are not required for activation of the Cpx response by overproduction of the outer-membrane lipoprotein NlpE or through mutation of the inner membrane protease HtpX. However, we found that respiration may play a role in activation of the Cpx response in E. coli lacking tolC, which encodes the outer membrane channel of multidrug efflux systems. We found that deletion of tolC activates the Cpx response through accumulation of the siderophores and that this decreases activity of the NADH-dependent aerobic electron transport chain. We provide evidence to suggest that enterobactin accumulation disrupts heme biosynthesis and that this may disrupt biogenesis of respiratory complexes. Finally, we have found that the Cpx response inhibits expression of genes involved in enterobactin biosynthesis. The results presented in this thesis make a significant contribution to the study of envelope biogenesis in gram-negative bacteria. We believe that activation of the Cpx response reduces expression of envelope-localized macromolecular protein complexes to prevent damage to the inner membrane. These data provide support for the hypothesis that the Cpx response functions to monitor and maintain the integrity of the inner membrane.

• Subjects / Keywords

  • Respiration
  • Electron Transport Chain
  • Escherichia coli
  • Envelope Stress Response

• Graduation date

  Fall 2017

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/R3707X48H

• License

  This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.