The Cpx stress response regulates turnover of respiratory chain proteins at the inner membrane of Escherichia coli
Creators: Tsviklist, Valeriya
Publisher: name: University of Alberta Library
Year: 2022

<p>The cell envelope of Gram-negative bacteria is a unique multilayered structure that protects them from the constantly changing and often times inhospitable environments, and  significantly contributes to their virulence. Having a complex structure, it requires an extensive regulatory network to monitor its assembly and stability. Escherichia coli possess an array of two-component signal transduction systems, some of which respond to cell envelope perturbations, including loss of proton-motive force, peptidoglycan stress, outer and inner membrane biogenesis defects. The Cpx envelope stress response is one of the major signaling pathways monitoring bacterial envelope integrity, activated both internally by excessive synthesis of membrane proteins and externally by a variety of environmental cues. The Cpx regulon is enriched with genes coding for protein folding and degrading factors, virulence determinants, large envelope-localized complexes and small regulatory RNAs. The Cpx response has been linked to a number of essential cellular processes, including iron sequestration, solute transport and cellular respiration. Transcriptional repression of the two electron transport chain complexes, NADH dehydrogenase I and cytochrome bo3, by the Cpx pathway has been demonstrated, however, there is evidence that additional regulatory mechanisms exist. The purpose of this thesis was to examine the interaction between Cpx-regulated protein folding and degrading factors and the respiratory complexes NADH dehydrogenase I and succinate dehydrogenase in Escherichia coli. Previously performed microarray analysis demonstrated that genes coding for succinate dehydrogenase complex were downregulated in the presence of the induced Cpx response. Here, we validate the microarray results and show that the succinate dehydrogenase complex has reduced activity in E. coli lacking the Cpx pathway and is most likely transcriptionally downregulated by the Cpx response.  Furthermore, we demonstrate that the stability of the NADH dehydrogenase I protein complex is lower in cells with a functional Cpx response, while in its absence, protein turnover is impaired. We provide evidence that Cpx-regulated envelope quality control factors are involved in the biogenesis and turnover of the respiratory complexes. Next, we demonstrate that the cellular need for Cpx-mediated stress adaptation increases when respiratory complexes are more prevalent or active, which is demonstrated by the growth defect of Cpx-deficient strains on media that requires a functional electron transport chain. Interestingly, deletion of several Cpx-regulated proteolytic factors and chaperones resulted in similar growth-deficient phenotypes. Finally, we have found that increased expression of the small regulatory RNA CpxQ, whose production is strongly upregulated upon Cpx induction, leads to stabilization or increased transcription of the sdhC transcript. Together, our results demonstrate that the Cpx two-component system has a broader function in surveillance of the cell envelope, regulating the abundance of large envelope protein complexes. It mounts a complex and balanced response to envelope damage, allowing effective recovery from the envelope stress and maintaining the cellular energy status of the cell. </p>