<i>In vivo</i> localizations of membrane stress controllers PspA and PspG in <i>Escherichia coli</i>

Engl, Christoph; Jovanović, Goran; Lloyd, Louise J.; Murray, Heath; Spitaler, Martin; Ying, Liming; Errington, Jeff; Buck, Martin

doi:10.1111/j.1365-2958.2009.06776.x

Cited by 62 publications

(94 citation statements)

References 49 publications

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“…Transcription from the promoters of both operons is 54 dependent and is activated by the transcriptional dual regulator PspF (39). PspA is believed to play a physical role in stabilizing membranes (24,80) and has been shown to suppress proton leakage of damaged liposomes composed of E. coli phospholipids (46).…”

Section: Resultsmentioning

confidence: 99%

Membrane Stresses Induced by Overproduction of Free Fatty Acids in Escherichia coli

Lennen

Kruziki

Kumar

et al. 2011

Appl Environ Microbiol

138

157

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Microbially produced fatty acids are potential precursors to high-energy-density biofuels, including alkanes and alkyl ethyl esters, by either catalytic conversion of free fatty acids (FFAs) or enzymatic conversion of acyl-acyl carrier protein or acyl-coenzyme A intermediates. Metabolic engineering efforts aimed at overproducing FFAs in Escherichia coli have achieved less than 30% of the maximum theoretical yield on the supplied carbon source. In this work, the viability, morphology, transcript levels, and protein levels of a strain of E. coli that overproduces medium-chain-length FFAs was compared to an engineered control strain. By early stationary phase, an 85% reduction in viable cell counts and exacerbated loss of inner membrane integrity were observed in the FFA-overproducing strain. These effects were enhanced in strains endogenously producing FFAs compared to strains exposed to exogenously fed FFAs. Under two sets of cultivation conditions, longchain unsaturated fatty acid content greatly increased, and the expression of genes and proteins required for unsaturated fatty acid biosynthesis were significantly decreased. Membrane stresses were further implicated by increased expression of genes and proteins of the phage shock response, the MarA/Rob/SoxS regulon, and the nuo and cyo operons of aerobic respiration. Gene deletion studies confirmed the importance of the phage shock proteins and Rob for maintaining cell viability; however, little to no change in FFA titer was observed after 24 h of cultivation. The results of this study serve as a baseline for future targeted attempts to improve FFA yields and titers in E. coli.Microbially derived free fatty acids (FFAs) are attractive intermediates for producing a wide range of high-energy-density biofuels from sustainable carbon sources, such as biomass (34). FFAs can be extracted from culture medium and catalytically converted to esters or alkanes (48, 55). Alternatively, enzymatic pathways exist for intracellular conversion to esters (42, 81), olefins (10, 59, 75), alkanes (78), or fatty aldehydes and fatty alcohols (22,81,82). These pathways can either be exploited in their native host or heterologously expressed in a genetically pliable microorganism (3). The physical and chemical properties of the resulting products are dependent on chain length and hydrophobicity; however, medium-chainlength (8-to 14-carbon) methyl esters, olefins, and alkanes exhibit many properties analogous to those of diesel and jet fuel and are therefore potential drop-in replacements (44,61).Several studies have demonstrated FFA overproduction in Escherichia coli (19,48,52,81,83). In each, the key strain modifications included overexpression of one or more cytosolic acyl-acyl carrier protein (ACP) thioesterases and deletion of fadD, or both fadD and fadE, which encode an acyl-coenzyme A (CoA) synthetase and acyl-CoA dehydrogenase, respectively. Overexpression of an acyl-ACP thioesterase depletes the level of acyl-ACP intermediates, which inhibit via feedback enzymes of fatty acid bi...

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Section: Resultsmentioning

confidence: 99%

Membrane Stresses Induced by Overproduction of Free Fatty Acids in Escherichia coli

Lennen

Kruziki

Kumar

et al. 2011

Appl Environ Microbiol

138

157

View full text Add to dashboard Cite

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“…These observations, along with those of other reports (Zhang and Sakamoto, 2013;Hennig et al, 2015), corroborate the important role of VIPP1 in photosynthetic membrane integrity. Movement associated with membrane stress has also been reported in PspA (Engl et al, 2009;Yamaguchi et al, 2013).…”

mentioning

confidence: 87%

VIPP1 Has a Disordered C-Terminal Tail Necessary for Protecting Photosynthetic Membranes against Stress

et al. 2016

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Integrity of biomembranes is vital to living organisms. In bacteria, PspA is considered to act as repairing damaged membrane by forming large supercomplexes in Arabidopsis (Arabidopsis thaliana). Vulnerable to oxidative stress, photosynthetic organisms also contain a PspA ortholog called VIPP1, which has an additional C-terminal tail (Vc). In this study, Vc was shown to coincide with an intrinsically disordered region, and the role of VIPP1 in membrane protection against stress was investigated. We visualized VIPP1 by fusing it to GFP (VIPP1-GFP that fully complemented lethal vipp1 mutations), and investigated its behavior in vivo with live imaging. The intrinsically disordered nature of Vc enabled VIPP1 to form what appeared to be functional particles along envelopes, whereas the deletion of Vc caused excessive association of the VIPP1 particles, preventing their active movement for membrane protection. Expression of VIPP1 lacking Vc complemented vipp1 mutation, but exhibited sensitivity to heat shock stress. Conversely, transgenic plants over-expressing VIPP1 showed enhanced tolerance against heat shock, suggesting that Vc negatively regulates VIPP1 particle association and acts in maintaining membrane integrity. Our data thus indicate that VIPP1 is involved in the maintenance of photosynthetic membranes. During evolution, chloroplasts have acquired enhanced tolerance against membrane stress by incorporating a disordered C-terminal tail into VIPP1.

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“…RSP28, RSP28A, RSP28B, RSP28F, RSP28AB, and RSP28ABF (Table 1). RSP28, RSP28A, RSP28B, RSP28F, RSP28AB, and RSP28ABF were constructed by transducing strains TB28, RS28A, RS28B, RS28F, RS28AB, and RS28ABF, respectively, to Amp r using a P1 lysate from strain MVA4 (46). TB28sc and RS28ABsc.…”

Section: Methodsmentioning

confidence: 99%

“…The membrane potential (⌬) was determined using a modified version of a previously described protocol (25,46) employing the JC-1 red/green dye-based assay. JC-1 (Life Technologies) was prepared as a stock of 5 mg/ml in dimethyl sulfoxide (DMSO) and stored at Ϫ20°C until use.…”

Section: Methodsmentioning

confidence: 99%

Multiple Envelope Stress Response Pathways Are Activated in an Escherichia coli Strain with Mutations in Two Members of the DedA Membrane Protein Family

2013

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We have reported that simultaneous deletion of two Escherichia coli genes, yqjA and yghB, encoding related and conserved inner membrane proteins belonging to the DedA protein family results in a number of intriguing phenotypes, including temperature sensitivity at 42°C, altered membrane lipid composition, and cell division defects. We sought to characterize these and other phenotypes in an effort to establish a function for this protein family in E. coli. Here, using reporter assays, we show that the major envelope stress response pathways Cpx, Psp, Bae, and Rcs are activated in strain BC202 (W3110; ⌬yqjA ⌬yghB) at the permissive growth temperature of 30°C. We previously demonstrated that 10 mM Mg 2؉ , 400 mM NaCl, and overexpression of tatABC are capable of restoring normal growth to BC202 at elevated growth temperatures. Deletion of the cpxR gene from BC202 results in the loss of the ability of these supplements to restore growth at 42°C. Additionally, we report that the membrane potential of BC202 is significantly reduced and that cell division and growth can be restored either by expression of the multidrug transporter MdfA from a multicopy plasmid or by growth at pH 6.0. Together, these results suggest that the DedA family proteins YqjA and YghB are required for general envelope maintenance and homeostasis of the proton motive force under a variety of growth conditions.

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In vivo localizations of membrane stress controllers PspA and PspG in Escherichia coli

Cited by 62 publications

References 49 publications

Membrane Stresses Induced by Overproduction of Free Fatty Acids in Escherichia coli

Membrane Stresses Induced by Overproduction of Free Fatty Acids in Escherichia coli

VIPP1 Has a Disordered C-Terminal Tail Necessary for Protecting Photosynthetic Membranes against Stress

Multiple Envelope Stress Response Pathways Are Activated in an Escherichia coli Strain with Mutations in Two Members of the DedA Membrane Protein Family

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