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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

Interaction study of the SpoIIS proteins using the BACTH system. Since fusions with SpoIIS proteins in both orientations were positive in some cases, only representative ones were selected. A strain expressing a pair of leucine zipper proteins, T25-Zip and T18-Zip, served as the positive control; the negative control was a strain expressing the pair T25-CIISA Bc and T18-Zip. Abbreviations: Bc, B. cereus; Bs, B. subtilis; CIISA, C-terminal domain of SpoIISA; IISB, SpoIISB; IISC, SpoIISC.
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Figure 3: Interaction study of the SpoIIS proteins using the BACTH system. Since fusions with SpoIIS proteins in both orientations were positive in some cases, only representative ones were selected. A strain expressing a pair of leucine zipper proteins, T25-Zip and T18-Zip, served as the positive control; the negative control was a strain expressing the pair T25-CIISA Bc and T18-Zip. Abbreviations: Bc, B. cereus; Bs, B. subtilis; CIISA, C-terminal domain of SpoIISA; IISB, SpoIISB; IISC, SpoIISC.

Mentions: The clearest evidence that B. subtilis SpoIISA and SpoIISB directly interact can be found in the crystal structure of the C-terminal domain of SpoIISA in complex with SpoIISB (Florek et al., 2011). To analyze the protein–protein interactions of the B. cereus SpoIIS proteins in vivo, we made use of the bacterial adenylate cyclase two hybrid system (Karimova et al., 1998). Like B. subtilis SpoIISA, B. cereus SpoIISA is predicted to be a membrane protein with three membrane-spanning segments. However, we decided to use only the cytoplasmic domains in this protein–protein interaction study, since the whole SpoIISA protein is toxic for E. coli as we have shown previously. We prepared fusions of the C-terminal domain of B. cereus SpoIISA, SpoIISB, and SpoIISC with the adenylate cyclase fragments T25 and T18. All possible interactions were tested and compared with those of similar SpoIIS fusion proteins from B. subtilis (Figure 3).


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)

Interaction study of the SpoIIS proteins using the BACTH system. Since fusions with SpoIIS proteins in both orientations were positive in some cases, only representative ones were selected. A strain expressing a pair of leucine zipper proteins, T25-Zip and T18-Zip, served as the positive control; the negative control was a strain expressing the pair T25-CIISA Bc and T18-Zip. Abbreviations: Bc, B. cereus; Bs, B. subtilis; CIISA, C-terminal domain of SpoIISA; IISB, SpoIISB; IISC, SpoIISC.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 3: Interaction study of the SpoIIS proteins using the BACTH system. Since fusions with SpoIIS proteins in both orientations were positive in some cases, only representative ones were selected. A strain expressing a pair of leucine zipper proteins, T25-Zip and T18-Zip, served as the positive control; the negative control was a strain expressing the pair T25-CIISA Bc and T18-Zip. Abbreviations: Bc, B. cereus; Bs, B. subtilis; CIISA, C-terminal domain of SpoIISA; IISB, SpoIISB; IISC, SpoIISC.
Mentions: The clearest evidence that B. subtilis SpoIISA and SpoIISB directly interact can be found in the crystal structure of the C-terminal domain of SpoIISA in complex with SpoIISB (Florek et al., 2011). To analyze the protein–protein interactions of the B. cereus SpoIIS proteins in vivo, we made use of the bacterial adenylate cyclase two hybrid system (Karimova et al., 1998). Like B. subtilis SpoIISA, B. cereus SpoIISA is predicted to be a membrane protein with three membrane-spanning segments. However, we decided to use only the cytoplasmic domains in this protein–protein interaction study, since the whole SpoIISA protein is toxic for E. coli as we have shown previously. We prepared fusions of the C-terminal domain of B. cereus SpoIISA, SpoIISB, and SpoIISC with the adenylate cyclase fragments T25 and T18. All possible interactions were tested and compared with those of similar SpoIIS fusion proteins from B. subtilis (Figure 3).

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