Limits...
Analysis of the Bacillus cereus SpoIIS antitoxin-toxin system reveals its three-component nature.

Melničáková J, Bečárová Z, Makroczyová J, Barák I - Front Microbiol (2015)

Bottom Line: In this work we describe the Bacillus cereus SpoIIS system which is a three-component system, harboring an additional gene spoIISC.Our results indicate that SpoIISC seems to be present not only in B. cereus but also in other Bacilli containing a SpoIIS toxin-antitoxin system.In addition, we show that B. cereus SpoIISA can form higher oligomers and we discuss the possible role of this multimerization for the protein's toxic function.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology, Slovak Academy of Sciences Bratislava, Slovakia.

ABSTRACT
Programmed cell death in bacteria is generally associated with two-component toxin-antitoxin systems. The SpoIIS toxin-antitoxin system, consisting of a membrane-bound SpoIISA toxin and a small, cytosolic antitoxin SpoIISB, was originally identified in Bacillus subtilis. In this work we describe the Bacillus cereus SpoIIS system which is a three-component system, harboring an additional gene spoIISC. Its protein product serves as an antitoxin, and similarly as SpoIISB, is able to bind SpoIISA and abolish its toxic effect. Our results indicate that SpoIISC seems to be present not only in B. cereus but also in other Bacilli containing a SpoIIS toxin-antitoxin system. In addition, we show that B. cereus SpoIISA can form higher oligomers and we discuss the possible role of this multimerization for the protein's toxic function.

No MeSH data available.


Related in: MedlinePlus

Kill/rescue assay in E. coli MM294. In order to test the ability of SpoIISC to act as an antitoxin for SpoIISA, SpoIIS proteins were expressed alone or in combination in E. coli cells. All results are mean values of three independent replicates and the bars represent 1 SD. The growth of E. coli cells expressing the SpoIISA toxin (circle) was inhibited while those cells expressing either the SpoIISAB complex (square) or the SpoIISAC complex (triangle) had wild-type growth (no marker). Arrows indicate the addition of 0.02% arabinose to induce expression. (A) The effect of the B. cereus SpoIIS proteins on the growth of E. coli MM294 cells. (B) The effect of the B. subtilis SpoIIS proteins on the growth of E. coli MM294 cells.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4526809&req=5

Figure 2: Kill/rescue assay in E. coli MM294. In order to test the ability of SpoIISC to act as an antitoxin for SpoIISA, SpoIIS proteins were expressed alone or in combination in E. coli cells. All results are mean values of three independent replicates and the bars represent 1 SD. The growth of E. coli cells expressing the SpoIISA toxin (circle) was inhibited while those cells expressing either the SpoIISAB complex (square) or the SpoIISAC complex (triangle) had wild-type growth (no marker). Arrows indicate the addition of 0.02% arabinose to induce expression. (A) The effect of the B. cereus SpoIIS proteins on the growth of E. coli MM294 cells. (B) The effect of the B. subtilis SpoIIS proteins on the growth of E. coli MM294 cells.

Mentions: To determine if B. cereus SpoIISB and SpoIISC are both able to neutralize the toxicity of B. cereus SpoIISA in E. coli, the corresponding genes spoIISAB and spoIISAC were cloned into pBAD24 vectors under the control of arabinose-inducible PBAD promoters to generate pBADIISAB Bc and pBADIISAC Bc. These plasmids were subsequently introduced into E. coli MM294 cells. The growth of these transformed cells, together with the control strains IB890 (E. coli MM294 / pBAD24) and IB926 (E. coli MM294/pBAD-BCIISA) (Florek et al., 2008), was monitored after the induction of protein expression. As found previously (Florek et al., 2008), the growth of E. coli cells expressing only B. cereus SpoIISA was inhibited. On the other hand, both SpoIISB and SpoIISC were able to neutralize the toxicity of SpoIISA: the growth curves of those strains which expressed both SpoIISA and either the SpoIISB antitoxin or SpoIISC were similar to that of the wild-type IB890 E. coli cells (Figure 2A). Because B. cereus SpoIISB and SpoIISC disturb SpoIISA toxicity when expressed in E. coli cells, it can be concluded that both spoIISB and spoIISC encode antitoxins and that they are likely to have similar functions as the antitoxins in B. subtilis. Indeed, an identical set of experiments using the B. subtilis genes rather than the B. cereus ones gives very similar results (Figure 2B).


Analysis of the Bacillus cereus SpoIIS antitoxin-toxin system reveals its three-component nature.

Melničáková J, Bečárová Z, Makroczyová J, Barák I - Front Microbiol (2015)

Kill/rescue assay in E. coli MM294. In order to test the ability of SpoIISC to act as an antitoxin for SpoIISA, SpoIIS proteins were expressed alone or in combination in E. coli cells. All results are mean values of three independent replicates and the bars represent 1 SD. The growth of E. coli cells expressing the SpoIISA toxin (circle) was inhibited while those cells expressing either the SpoIISAB complex (square) or the SpoIISAC complex (triangle) had wild-type growth (no marker). Arrows indicate the addition of 0.02% arabinose to induce expression. (A) The effect of the B. cereus SpoIIS proteins on the growth of E. coli MM294 cells. (B) The effect of the B. subtilis SpoIIS proteins on the growth of E. coli MM294 cells.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526809&req=5

Figure 2: Kill/rescue assay in E. coli MM294. In order to test the ability of SpoIISC to act as an antitoxin for SpoIISA, SpoIIS proteins were expressed alone or in combination in E. coli cells. All results are mean values of three independent replicates and the bars represent 1 SD. The growth of E. coli cells expressing the SpoIISA toxin (circle) was inhibited while those cells expressing either the SpoIISAB complex (square) or the SpoIISAC complex (triangle) had wild-type growth (no marker). Arrows indicate the addition of 0.02% arabinose to induce expression. (A) The effect of the B. cereus SpoIIS proteins on the growth of E. coli MM294 cells. (B) The effect of the B. subtilis SpoIIS proteins on the growth of E. coli MM294 cells.
Mentions: To determine if B. cereus SpoIISB and SpoIISC are both able to neutralize the toxicity of B. cereus SpoIISA in E. coli, the corresponding genes spoIISAB and spoIISAC were cloned into pBAD24 vectors under the control of arabinose-inducible PBAD promoters to generate pBADIISAB Bc and pBADIISAC Bc. These plasmids were subsequently introduced into E. coli MM294 cells. The growth of these transformed cells, together with the control strains IB890 (E. coli MM294 / pBAD24) and IB926 (E. coli MM294/pBAD-BCIISA) (Florek et al., 2008), was monitored after the induction of protein expression. As found previously (Florek et al., 2008), the growth of E. coli cells expressing only B. cereus SpoIISA was inhibited. On the other hand, both SpoIISB and SpoIISC were able to neutralize the toxicity of SpoIISA: the growth curves of those strains which expressed both SpoIISA and either the SpoIISB antitoxin or SpoIISC were similar to that of the wild-type IB890 E. coli cells (Figure 2A). Because B. cereus SpoIISB and SpoIISC disturb SpoIISA toxicity when expressed in E. coli cells, it can be concluded that both spoIISB and spoIISC encode antitoxins and that they are likely to have similar functions as the antitoxins in B. subtilis. Indeed, an identical set of experiments using the B. subtilis genes rather than the B. cereus ones gives very similar results (Figure 2B).

Bottom Line: In this work we describe the Bacillus cereus SpoIIS system which is a three-component system, harboring an additional gene spoIISC.Our results indicate that SpoIISC seems to be present not only in B. cereus but also in other Bacilli containing a SpoIIS toxin-antitoxin system.In addition, we show that B. cereus SpoIISA can form higher oligomers and we discuss the possible role of this multimerization for the protein's toxic function.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology, Slovak Academy of Sciences Bratislava, Slovakia.

ABSTRACT
Programmed cell death in bacteria is generally associated with two-component toxin-antitoxin systems. The SpoIIS toxin-antitoxin system, consisting of a membrane-bound SpoIISA toxin and a small, cytosolic antitoxin SpoIISB, was originally identified in Bacillus subtilis. In this work we describe the Bacillus cereus SpoIIS system which is a three-component system, harboring an additional gene spoIISC. Its protein product serves as an antitoxin, and similarly as SpoIISB, is able to bind SpoIISA and abolish its toxic effect. Our results indicate that SpoIISC seems to be present not only in B. cereus but also in other Bacilli containing a SpoIIS toxin-antitoxin system. In addition, we show that B. cereus SpoIISA can form higher oligomers and we discuss the possible role of this multimerization for the protein's toxic function.

No MeSH data available.


Related in: MedlinePlus