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Two programmed cell death systems in Escherichia coli: an apoptotic-like death is inhibited by the mazEF-mediated death pathway.

Erental A, Sharon I, Engelberg-Kulka H - PLoS Biol. (2012)

Bottom Line: The mazEF-mediated pathway reduces recA mRNA levels.Based on these results, we offer a molecular model for the maintenance of an altruistic characteristic in cell populations.In our model, the ALD pathway is inhibited by the altruistic EDF-mazEF-mediated death pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Molecular Genetics, Institute for Medical Research Israel-Canada, The Hebrew University-Hadassah Medical School, Jerusalem, Israel.

ABSTRACT
In eukaryotes, the classical form of programmed cell death (PCD) is apoptosis, which has as its specific characteristics DNA fragmentation and membrane depolarization. In Escherichia coli a different PCD system has been reported. It is mediated by the toxin-antitoxin system module mazEF. The E. coli mazEF module is one of the most thoroughly studied toxin-antitoxin systems. mazF encodes a stable toxin, MazF, and mazE encodes a labile antitoxin, MazE, which prevents the lethal effect of MazF. mazEF-mediated cell death is a population phenomenon requiring the quorum-sensing pentapeptide NNWNN designated Extracellular Death Factor (EDF). mazEF is triggered by several stressful conditions, including severe damage to the DNA. Here, using confocal microscopy and FACS analysis, we show that under conditions of severe DNA damage, the triggered mazEF-mediated cell death pathway leads to the inhibition of a second cell death pathway. The latter is an apoptotic-like death (ALD); ALD is mediated by recA and lexA. The mazEF-mediated pathway reduces recA mRNA levels. Based on these results, we offer a molecular model for the maintenance of an altruistic characteristic in cell populations. In our model, the ALD pathway is inhibited by the altruistic EDF-mazEF-mediated death pathway.

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The mazEF pathway reduces the levels of recA mRNA.(A) E. coli cells of strain MC4100relA+, its ΔmazEF derivative, or ΔmazEF carrying plasmid pKKmazEF were grown and treated with NA (100 µg/ml) or left without treatment (n.t) as in Figure 1. Then RNA was extracted from the cells, and real-time PCR was performed to quantify recA mRNA levels. The indicated values are relative to recA RNA levels in untreated MC4100relA+ cells. (B) E. coli MC4100relA+ΔyfiD or E. coli MC4100relA+ΔyfiD cells carrying the inducible plasmid pQE-yfiD were grown and treated with NA as in (A). To induce the expression of yfiD, 1 mM of IPTG was added before the addition of NA. The indicated values are relative to recA RNA levels in untreated MC4100relA+ΔyfiD cells. (C) E. coli MG1655 cells were grown and treated with NA (100 µg/ml) or left without treatment as in (A). To test the effect of EDF on this strain, a synthetic EDF (50 ng/ml) or iEDF (50 ng/ml) (NNGNN) was added to the cells before adding the NA. The indicated values are relative to recA RNA levels in untreated MG1655 cells. All experiments were performed in triplicate, and a typical experiment out of three is shown. Error bars indicate standard deviation.
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pbio-1001281-g006: The mazEF pathway reduces the levels of recA mRNA.(A) E. coli cells of strain MC4100relA+, its ΔmazEF derivative, or ΔmazEF carrying plasmid pKKmazEF were grown and treated with NA (100 µg/ml) or left without treatment (n.t) as in Figure 1. Then RNA was extracted from the cells, and real-time PCR was performed to quantify recA mRNA levels. The indicated values are relative to recA RNA levels in untreated MC4100relA+ cells. (B) E. coli MC4100relA+ΔyfiD or E. coli MC4100relA+ΔyfiD cells carrying the inducible plasmid pQE-yfiD were grown and treated with NA as in (A). To induce the expression of yfiD, 1 mM of IPTG was added before the addition of NA. The indicated values are relative to recA RNA levels in untreated MC4100relA+ΔyfiD cells. (C) E. coli MG1655 cells were grown and treated with NA (100 µg/ml) or left without treatment as in (A). To test the effect of EDF on this strain, a synthetic EDF (50 ng/ml) or iEDF (50 ng/ml) (NNGNN) was added to the cells before adding the NA. The indicated values are relative to recA RNA levels in untreated MG1655 cells. All experiments were performed in triplicate, and a typical experiment out of three is shown. Error bars indicate standard deviation.

Mentions: Here we asked at what stage is the ALD pathway inhibited by the EDF-mazEF-mediated pathway? Since we have found that the ALD pathway is recA-dependent (Figure 4), here we examined whether the EDF-mazEF pathway affects levels of recA mRNA. As shown, the addition of NA to strain MC4100relA+ did not increase relative recA mRNA levels (Figure 6A). In contrast, these mRNA levels were significantly increased (about 40-fold) by the addition of NA to its ΔmazEF derivative (Figure 6A). Moreover, when ΔmazEF strains harbored plasmid pKKmazEF, which expresses mazEF genes constitutively, the relative mRNA levels of recA were decreased significantly in the presence of NA (Figure 6A). Since we have previously shown that yfiD acts downstream to MazF [26],[27], we here asked whether yfiD also reduces the levels of recA mRNA. As shown, indeed, the deletion of yfiD significantly elevated the level of recA mRNA in the presence of NA (Figure 6B). Moreover, when these ΔyfiD strains harbored plasmid pQE-yfiD, which bears an IPTG-inducible yfiD gene, and in the presence of 1 mM IPTG, mRNA levels of recA were significantly decreased in the presence of NA (Figure 6B). These results suggest that it is not MazF itself that directly reduces the mRNA levels of recA, but rather the product of one of its downstream genes. In addition, we also asked whether EDF is involved in the reduction of recA mRNA levels. To this aim, we used the E. coli strain MG1655, which is impaired in EDF production [29]. Indeed, the addition of 50 ng/ml synthetic EDF to the MG1655 strain significantly decreased recA mRNA levels in the presence of NA, whereas the addition of iEDF (NNGNN) did not affect the levels of this mRNA (Figure 6C).


Two programmed cell death systems in Escherichia coli: an apoptotic-like death is inhibited by the mazEF-mediated death pathway.

Erental A, Sharon I, Engelberg-Kulka H - PLoS Biol. (2012)

The mazEF pathway reduces the levels of recA mRNA.(A) E. coli cells of strain MC4100relA+, its ΔmazEF derivative, or ΔmazEF carrying plasmid pKKmazEF were grown and treated with NA (100 µg/ml) or left without treatment (n.t) as in Figure 1. Then RNA was extracted from the cells, and real-time PCR was performed to quantify recA mRNA levels. The indicated values are relative to recA RNA levels in untreated MC4100relA+ cells. (B) E. coli MC4100relA+ΔyfiD or E. coli MC4100relA+ΔyfiD cells carrying the inducible plasmid pQE-yfiD were grown and treated with NA as in (A). To induce the expression of yfiD, 1 mM of IPTG was added before the addition of NA. The indicated values are relative to recA RNA levels in untreated MC4100relA+ΔyfiD cells. (C) E. coli MG1655 cells were grown and treated with NA (100 µg/ml) or left without treatment as in (A). To test the effect of EDF on this strain, a synthetic EDF (50 ng/ml) or iEDF (50 ng/ml) (NNGNN) was added to the cells before adding the NA. The indicated values are relative to recA RNA levels in untreated MG1655 cells. All experiments were performed in triplicate, and a typical experiment out of three is shown. Error bars indicate standard deviation.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3295820&req=5

pbio-1001281-g006: The mazEF pathway reduces the levels of recA mRNA.(A) E. coli cells of strain MC4100relA+, its ΔmazEF derivative, or ΔmazEF carrying plasmid pKKmazEF were grown and treated with NA (100 µg/ml) or left without treatment (n.t) as in Figure 1. Then RNA was extracted from the cells, and real-time PCR was performed to quantify recA mRNA levels. The indicated values are relative to recA RNA levels in untreated MC4100relA+ cells. (B) E. coli MC4100relA+ΔyfiD or E. coli MC4100relA+ΔyfiD cells carrying the inducible plasmid pQE-yfiD were grown and treated with NA as in (A). To induce the expression of yfiD, 1 mM of IPTG was added before the addition of NA. The indicated values are relative to recA RNA levels in untreated MC4100relA+ΔyfiD cells. (C) E. coli MG1655 cells were grown and treated with NA (100 µg/ml) or left without treatment as in (A). To test the effect of EDF on this strain, a synthetic EDF (50 ng/ml) or iEDF (50 ng/ml) (NNGNN) was added to the cells before adding the NA. The indicated values are relative to recA RNA levels in untreated MG1655 cells. All experiments were performed in triplicate, and a typical experiment out of three is shown. Error bars indicate standard deviation.
Mentions: Here we asked at what stage is the ALD pathway inhibited by the EDF-mazEF-mediated pathway? Since we have found that the ALD pathway is recA-dependent (Figure 4), here we examined whether the EDF-mazEF pathway affects levels of recA mRNA. As shown, the addition of NA to strain MC4100relA+ did not increase relative recA mRNA levels (Figure 6A). In contrast, these mRNA levels were significantly increased (about 40-fold) by the addition of NA to its ΔmazEF derivative (Figure 6A). Moreover, when ΔmazEF strains harbored plasmid pKKmazEF, which expresses mazEF genes constitutively, the relative mRNA levels of recA were decreased significantly in the presence of NA (Figure 6A). Since we have previously shown that yfiD acts downstream to MazF [26],[27], we here asked whether yfiD also reduces the levels of recA mRNA. As shown, indeed, the deletion of yfiD significantly elevated the level of recA mRNA in the presence of NA (Figure 6B). Moreover, when these ΔyfiD strains harbored plasmid pQE-yfiD, which bears an IPTG-inducible yfiD gene, and in the presence of 1 mM IPTG, mRNA levels of recA were significantly decreased in the presence of NA (Figure 6B). These results suggest that it is not MazF itself that directly reduces the mRNA levels of recA, but rather the product of one of its downstream genes. In addition, we also asked whether EDF is involved in the reduction of recA mRNA levels. To this aim, we used the E. coli strain MG1655, which is impaired in EDF production [29]. Indeed, the addition of 50 ng/ml synthetic EDF to the MG1655 strain significantly decreased recA mRNA levels in the presence of NA, whereas the addition of iEDF (NNGNN) did not affect the levels of this mRNA (Figure 6C).

Bottom Line: The mazEF-mediated pathway reduces recA mRNA levels.Based on these results, we offer a molecular model for the maintenance of an altruistic characteristic in cell populations.In our model, the ALD pathway is inhibited by the altruistic EDF-mazEF-mediated death pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Molecular Genetics, Institute for Medical Research Israel-Canada, The Hebrew University-Hadassah Medical School, Jerusalem, Israel.

ABSTRACT
In eukaryotes, the classical form of programmed cell death (PCD) is apoptosis, which has as its specific characteristics DNA fragmentation and membrane depolarization. In Escherichia coli a different PCD system has been reported. It is mediated by the toxin-antitoxin system module mazEF. The E. coli mazEF module is one of the most thoroughly studied toxin-antitoxin systems. mazF encodes a stable toxin, MazF, and mazE encodes a labile antitoxin, MazE, which prevents the lethal effect of MazF. mazEF-mediated cell death is a population phenomenon requiring the quorum-sensing pentapeptide NNWNN designated Extracellular Death Factor (EDF). mazEF is triggered by several stressful conditions, including severe damage to the DNA. Here, using confocal microscopy and FACS analysis, we show that under conditions of severe DNA damage, the triggered mazEF-mediated cell death pathway leads to the inhibition of a second cell death pathway. The latter is an apoptotic-like death (ALD); ALD is mediated by recA and lexA. The mazEF-mediated pathway reduces recA mRNA levels. Based on these results, we offer a molecular model for the maintenance of an altruistic characteristic in cell populations. In our model, the ALD pathway is inhibited by the altruistic EDF-mazEF-mediated death pathway.

Show MeSH
Related in: MedlinePlus