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The extracytoplasmic stress factor, sigmaE, is required to maintain cell envelope integrity in Escherichia coli.

Hayden JD, Ades SE - PLoS ONE (2008)

Bottom Line: Many cells lyse and some develop blebs containing cytoplasmic material along their sides.To better understand the connection between transcription by sigma(E) and cell envelope integrity, we identified two multicopy suppressors of the essentiality of sigma(E), ptsN and yhbW. yhbW is a gene of unknown function, while ptsN is a member of the sigma(E) regulon.Overexpression of ptsN lowers the basal level of multiple envelope stress responses, but not that of a cytoplasmic stress response.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, USA.

ABSTRACT
Extracytoplasmic function or ECF sigma factors are the most abundant class of alternative sigma factors in bacteria. Members of the rpoE subclass of ECF sigma factors are implicated in sensing stress in the cell envelope of Gram-negative bacteria and are required for virulence in many pathogens. The best-studied member of this family is rpoE from Escherichia coli, encoding the sigma(E) protein. sigma(E) has been well studied for its role in combating extracytoplasmic stress, and the members of its regulon have been largely defined. sigma(E) is required for viability of E. coli, yet none of the studies to date explain why sigma(E) is essential in seemingly unstressed cells. In this work we investigate the essential role of sigma(E) in E. coli by analyzing the phenotypes associated with loss of sigma(E) activity and isolating suppressors that allow cells to live in the absence of sigma(E). We demonstrate that when sigma(E) is inhibited, cell envelope stress increases and envelope integrity is lost. Many cells lyse and some develop blebs containing cytoplasmic material along their sides. To better understand the connection between transcription by sigma(E) and cell envelope integrity, we identified two multicopy suppressors of the essentiality of sigma(E), ptsN and yhbW. yhbW is a gene of unknown function, while ptsN is a member of the sigma(E) regulon. Overexpression of ptsN lowers the basal level of multiple envelope stress responses, but not that of a cytoplasmic stress response. Our results are consistent with a model in which overexpression of ptsN reduces stress in the cell envelope, thereby promoting survival in the absence of sigma(E).

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The σE-dependent extracytoplasmic stress response.σE is held at the membrane by the antisigma factor RseA. RseB binds to the periplasmic domain of RseA and protects RseA from proteolysis. Unfolded OMPs activate the protease DegS, which cleaves the periplasmic domain of RseA. The partially degraded RseA is now a substrate for RseP. RseP cleaves RseA further, releasing the cytoplasmic domain of RseA bound to σE. This remaining domain of RseA is degraded by ClpXP thereby freeing σE to interact with RNA polymerase and direct transcription of its regulon. The major classes of known genes in the σE regulon are indicated. The alarmone ppGpp and protein DksA can activate σE-dependent transcription once σE is released from RseA, but are not shown for clarity. (IM inner membrane, OM outer membrane).
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pone-0001573-g001: The σE-dependent extracytoplasmic stress response.σE is held at the membrane by the antisigma factor RseA. RseB binds to the periplasmic domain of RseA and protects RseA from proteolysis. Unfolded OMPs activate the protease DegS, which cleaves the periplasmic domain of RseA. The partially degraded RseA is now a substrate for RseP. RseP cleaves RseA further, releasing the cytoplasmic domain of RseA bound to σE. This remaining domain of RseA is degraded by ClpXP thereby freeing σE to interact with RNA polymerase and direct transcription of its regulon. The major classes of known genes in the σE regulon are indicated. The alarmone ppGpp and protein DksA can activate σE-dependent transcription once σE is released from RseA, but are not shown for clarity. (IM inner membrane, OM outer membrane).

Mentions: σE is activated by stresses that interfere with the folding of outer membrane proteins (OMPs) such as heat shock, overexpression of OMP genes, and mutations in genes encoding chaperones required for OMP folding [11]–[14]. In unstressed cells, σE activity is low because σE is sequestered at the inner membrane by the antisigma factor RseA (Fig. 1) [15], [16]. RseA is a single-pass inner membrane protein that binds to σE and prevents σE from interacting with RNA polymerase [15]–[17]. During envelope stress, RseA is degraded in response to unfolded porins by the sequential action of two inner membrane proteases, DegS and RseP, followed by the cytoplasmic protease ClpXP (Fig. 1) [18]–[22]. A periplasmic protein, RseB, binds to the periplasmic domain of RseA and enhances inhibition by RseA, protecting it from proteolysis (Fig. 1) [15], [16], [18], [23]. Continual degradation of RseA is required to provide the cell with sufficient free σE to support viability, and deletion of either degS or rseP is toxic due to the stabilization of RseA and consequent sequestration of σE [19], [20], [23]. σE can also be activated independently of the RseA-dependent stress-signaling pathway by the cytoplasmic alarmone ppGpp, whose levels change in response to nutrient availability [24].


The extracytoplasmic stress factor, sigmaE, is required to maintain cell envelope integrity in Escherichia coli.

Hayden JD, Ades SE - PLoS ONE (2008)

The σE-dependent extracytoplasmic stress response.σE is held at the membrane by the antisigma factor RseA. RseB binds to the periplasmic domain of RseA and protects RseA from proteolysis. Unfolded OMPs activate the protease DegS, which cleaves the periplasmic domain of RseA. The partially degraded RseA is now a substrate for RseP. RseP cleaves RseA further, releasing the cytoplasmic domain of RseA bound to σE. This remaining domain of RseA is degraded by ClpXP thereby freeing σE to interact with RNA polymerase and direct transcription of its regulon. The major classes of known genes in the σE regulon are indicated. The alarmone ppGpp and protein DksA can activate σE-dependent transcription once σE is released from RseA, but are not shown for clarity. (IM inner membrane, OM outer membrane).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2215328&req=5

pone-0001573-g001: The σE-dependent extracytoplasmic stress response.σE is held at the membrane by the antisigma factor RseA. RseB binds to the periplasmic domain of RseA and protects RseA from proteolysis. Unfolded OMPs activate the protease DegS, which cleaves the periplasmic domain of RseA. The partially degraded RseA is now a substrate for RseP. RseP cleaves RseA further, releasing the cytoplasmic domain of RseA bound to σE. This remaining domain of RseA is degraded by ClpXP thereby freeing σE to interact with RNA polymerase and direct transcription of its regulon. The major classes of known genes in the σE regulon are indicated. The alarmone ppGpp and protein DksA can activate σE-dependent transcription once σE is released from RseA, but are not shown for clarity. (IM inner membrane, OM outer membrane).
Mentions: σE is activated by stresses that interfere with the folding of outer membrane proteins (OMPs) such as heat shock, overexpression of OMP genes, and mutations in genes encoding chaperones required for OMP folding [11]–[14]. In unstressed cells, σE activity is low because σE is sequestered at the inner membrane by the antisigma factor RseA (Fig. 1) [15], [16]. RseA is a single-pass inner membrane protein that binds to σE and prevents σE from interacting with RNA polymerase [15]–[17]. During envelope stress, RseA is degraded in response to unfolded porins by the sequential action of two inner membrane proteases, DegS and RseP, followed by the cytoplasmic protease ClpXP (Fig. 1) [18]–[22]. A periplasmic protein, RseB, binds to the periplasmic domain of RseA and enhances inhibition by RseA, protecting it from proteolysis (Fig. 1) [15], [16], [18], [23]. Continual degradation of RseA is required to provide the cell with sufficient free σE to support viability, and deletion of either degS or rseP is toxic due to the stabilization of RseA and consequent sequestration of σE [19], [20], [23]. σE can also be activated independently of the RseA-dependent stress-signaling pathway by the cytoplasmic alarmone ppGpp, whose levels change in response to nutrient availability [24].

Bottom Line: Many cells lyse and some develop blebs containing cytoplasmic material along their sides.To better understand the connection between transcription by sigma(E) and cell envelope integrity, we identified two multicopy suppressors of the essentiality of sigma(E), ptsN and yhbW. yhbW is a gene of unknown function, while ptsN is a member of the sigma(E) regulon.Overexpression of ptsN lowers the basal level of multiple envelope stress responses, but not that of a cytoplasmic stress response.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, USA.

ABSTRACT
Extracytoplasmic function or ECF sigma factors are the most abundant class of alternative sigma factors in bacteria. Members of the rpoE subclass of ECF sigma factors are implicated in sensing stress in the cell envelope of Gram-negative bacteria and are required for virulence in many pathogens. The best-studied member of this family is rpoE from Escherichia coli, encoding the sigma(E) protein. sigma(E) has been well studied for its role in combating extracytoplasmic stress, and the members of its regulon have been largely defined. sigma(E) is required for viability of E. coli, yet none of the studies to date explain why sigma(E) is essential in seemingly unstressed cells. In this work we investigate the essential role of sigma(E) in E. coli by analyzing the phenotypes associated with loss of sigma(E) activity and isolating suppressors that allow cells to live in the absence of sigma(E). We demonstrate that when sigma(E) is inhibited, cell envelope stress increases and envelope integrity is lost. Many cells lyse and some develop blebs containing cytoplasmic material along their sides. To better understand the connection between transcription by sigma(E) and cell envelope integrity, we identified two multicopy suppressors of the essentiality of sigma(E), ptsN and yhbW. yhbW is a gene of unknown function, while ptsN is a member of the sigma(E) regulon. Overexpression of ptsN lowers the basal level of multiple envelope stress responses, but not that of a cytoplasmic stress response. Our results are consistent with a model in which overexpression of ptsN reduces stress in the cell envelope, thereby promoting survival in the absence of sigma(E).

Show MeSH
Related in: MedlinePlus