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Identification of the minimal cytolytic unit for streptolysin S and an expansion of the toxin family.

Molloy EM, Casjens SR, Cox CL, Maxson T, Ethridge NA, Margos G, Fingerle V, Mitchell DA - BMC Microbiol. (2015)

Bottom Line: Among the Spirochaete clusters, naturally truncated SLS-like precursors were found in the genomes of three Lyme disease-causing Borrelia burgdorferi sensu lato (Bbsl) strains.Interestingly, this region is found in all characterized TOMM cytolysins, as well as the novel putative TOMM cytolysins we discovered.In addition to providing new insight into the structure-activity relationships of SLS, this study greatly expands the cytolysin group of TOMMs.

View Article: PubMed Central - PubMed

Affiliation: Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL, 61801, USA. emolloy@illinois.edu.

ABSTRACT

Background: Streptolysin S (SLS) is a cytolytic virulence factor produced by the human pathogen Streptococcus pyogenes and other Streptococcus species. Related "SLS-like" toxins have been characterized in select strains of Clostridium and Listeria, with homologous clusters bioinformatically identified in a variety of other species. SLS is a member of the thiazole/oxazole-modified microcin (TOMM) family of natural products. The structure of SLS has yet to be deciphered and many questions remain regarding its structure-activity relationships.

Results: In this work, we assessed the hemolytic activity of a series of C-terminally truncated SLS peptides expressed in SLS-deficient S. pyogenes. Our data indicate that while the N-terminal poly-heterocyclizable (NPH) region of SLS substantially contributes to its bioactivity, the variable C-terminal region of the toxin is largely dispensable. Through genome mining we identified additional SLS-like clusters in diverse Firmicutes, Spirochaetes and Actinobacteria. Among the Spirochaete clusters, naturally truncated SLS-like precursors were found in the genomes of three Lyme disease-causing Borrelia burgdorferi sensu lato (Bbsl) strains. Although unable to restore hemolysis in SLS-deficient S. pyogenes, a Bbsl SLS-like precursor peptide was converted to a cytolysin using purified SLS biosynthetic enzymes. A PCR-based screen demonstrated that SLS-like clusters are substantially more prevalent in Bbsl than inferred from publicly available genome sequences.

Conclusions: The mutagenesis data described herein indicate that the minimal cytolytic unit of SLS encompasses the NPH region of the core peptide. Interestingly, this region is found in all characterized TOMM cytolysins, as well as the novel putative TOMM cytolysins we discovered. We propose that this conserved region represents the defining feature of the SLS-like TOMM family. We demonstrate the cytolytic potential of a Bbsl SLS-like precursor peptide, which has a core region of similar length to the SLS minimal cytolytic unit, when modified with purified SLS biosynthetic enzymes. As such, we speculate that some Borrelia have the potential to produce a TOMM cytolysin, although the biological significance of this finding remains to be determined. In addition to providing new insight into the structure-activity relationships of SLS, this study greatly expands the cytolysin group of TOMMs.

No MeSH data available.


Related in: MedlinePlus

Phylogenetic analysis of the TOMM cytolysin family. A maximum-likelihood tree based on the D protein (listed in Additional file 4: Table S2) for the majority of known and putative producers of cytolytic TOMMs (as of December 2014). This phylogenetic data is deposited in the Dryad Digital Repository, doi:10.5061/dryad.d4863. Clades are color-coded based on the originating genus for each SLS-like toxin, as represented in the legend. The relevant phyla are given in parentheses
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Fig3: Phylogenetic analysis of the TOMM cytolysin family. A maximum-likelihood tree based on the D protein (listed in Additional file 4: Table S2) for the majority of known and putative producers of cytolytic TOMMs (as of December 2014). This phylogenetic data is deposited in the Dryad Digital Repository, doi:10.5061/dryad.d4863. Clades are color-coded based on the originating genus for each SLS-like toxin, as represented in the legend. The relevant phyla are given in parentheses

Mentions: Given that the number of bioinformatically identifiable TOMM biosynthetic clusters more than quadrupled since 2011 [8, 25], we sought to determine if naturally truncated precursor peptides are present in any novel SLS-like clusters. Our genome mining effort revealed TOMM clusters encoding SagB, −C and –D homologs in the vicinity of SagA homologs in Firmicutes genera not previously known to possess such clusters (i.e. Bacillus, Lactobacillus, Enterococcus, Exiguobacterium, Oenococcus and Virgibacillus) (Additional file 2: Figure S2A and Additional file 3: Table S1). Interestingly, a number of Actinobacteria (Mobiluncus and Propionibacterium) and Spirochaetes (Borrelia and Brachyspira) encode SLS-like clusters (Additional file 2: Figure S2A and Additional file 3: Table S1). The Brachyspira TOMM clusters were independently noted in a recent publication [26]. We hypothesized that the newly-annotated SLS-like precursors (Additional file 2: Figure S2B) represent novel TOMM cytolysins since the sequence of a TOMM precursor dictates the function of the natural product after posttranslational modification [8, 6]. Moreover, while the novel SagA-like precursor peptides were of variable length, all contained the aforementioned NPH region. Our identification of these putative SLS-like cytolysins greatly expands the TOMM cytolysin family (Fig. 3 and Additional file 4: Table S2) and suggests that the TOMM cytolysins are not confined to Firmicutes.Fig. 3


Identification of the minimal cytolytic unit for streptolysin S and an expansion of the toxin family.

Molloy EM, Casjens SR, Cox CL, Maxson T, Ethridge NA, Margos G, Fingerle V, Mitchell DA - BMC Microbiol. (2015)

Phylogenetic analysis of the TOMM cytolysin family. A maximum-likelihood tree based on the D protein (listed in Additional file 4: Table S2) for the majority of known and putative producers of cytolytic TOMMs (as of December 2014). This phylogenetic data is deposited in the Dryad Digital Repository, doi:10.5061/dryad.d4863. Clades are color-coded based on the originating genus for each SLS-like toxin, as represented in the legend. The relevant phyla are given in parentheses
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4513790&req=5

Fig3: Phylogenetic analysis of the TOMM cytolysin family. A maximum-likelihood tree based on the D protein (listed in Additional file 4: Table S2) for the majority of known and putative producers of cytolytic TOMMs (as of December 2014). This phylogenetic data is deposited in the Dryad Digital Repository, doi:10.5061/dryad.d4863. Clades are color-coded based on the originating genus for each SLS-like toxin, as represented in the legend. The relevant phyla are given in parentheses
Mentions: Given that the number of bioinformatically identifiable TOMM biosynthetic clusters more than quadrupled since 2011 [8, 25], we sought to determine if naturally truncated precursor peptides are present in any novel SLS-like clusters. Our genome mining effort revealed TOMM clusters encoding SagB, −C and –D homologs in the vicinity of SagA homologs in Firmicutes genera not previously known to possess such clusters (i.e. Bacillus, Lactobacillus, Enterococcus, Exiguobacterium, Oenococcus and Virgibacillus) (Additional file 2: Figure S2A and Additional file 3: Table S1). Interestingly, a number of Actinobacteria (Mobiluncus and Propionibacterium) and Spirochaetes (Borrelia and Brachyspira) encode SLS-like clusters (Additional file 2: Figure S2A and Additional file 3: Table S1). The Brachyspira TOMM clusters were independently noted in a recent publication [26]. We hypothesized that the newly-annotated SLS-like precursors (Additional file 2: Figure S2B) represent novel TOMM cytolysins since the sequence of a TOMM precursor dictates the function of the natural product after posttranslational modification [8, 6]. Moreover, while the novel SagA-like precursor peptides were of variable length, all contained the aforementioned NPH region. Our identification of these putative SLS-like cytolysins greatly expands the TOMM cytolysin family (Fig. 3 and Additional file 4: Table S2) and suggests that the TOMM cytolysins are not confined to Firmicutes.Fig. 3

Bottom Line: Among the Spirochaete clusters, naturally truncated SLS-like precursors were found in the genomes of three Lyme disease-causing Borrelia burgdorferi sensu lato (Bbsl) strains.Interestingly, this region is found in all characterized TOMM cytolysins, as well as the novel putative TOMM cytolysins we discovered.In addition to providing new insight into the structure-activity relationships of SLS, this study greatly expands the cytolysin group of TOMMs.

View Article: PubMed Central - PubMed

Affiliation: Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL, 61801, USA. emolloy@illinois.edu.

ABSTRACT

Background: Streptolysin S (SLS) is a cytolytic virulence factor produced by the human pathogen Streptococcus pyogenes and other Streptococcus species. Related "SLS-like" toxins have been characterized in select strains of Clostridium and Listeria, with homologous clusters bioinformatically identified in a variety of other species. SLS is a member of the thiazole/oxazole-modified microcin (TOMM) family of natural products. The structure of SLS has yet to be deciphered and many questions remain regarding its structure-activity relationships.

Results: In this work, we assessed the hemolytic activity of a series of C-terminally truncated SLS peptides expressed in SLS-deficient S. pyogenes. Our data indicate that while the N-terminal poly-heterocyclizable (NPH) region of SLS substantially contributes to its bioactivity, the variable C-terminal region of the toxin is largely dispensable. Through genome mining we identified additional SLS-like clusters in diverse Firmicutes, Spirochaetes and Actinobacteria. Among the Spirochaete clusters, naturally truncated SLS-like precursors were found in the genomes of three Lyme disease-causing Borrelia burgdorferi sensu lato (Bbsl) strains. Although unable to restore hemolysis in SLS-deficient S. pyogenes, a Bbsl SLS-like precursor peptide was converted to a cytolysin using purified SLS biosynthetic enzymes. A PCR-based screen demonstrated that SLS-like clusters are substantially more prevalent in Bbsl than inferred from publicly available genome sequences.

Conclusions: The mutagenesis data described herein indicate that the minimal cytolytic unit of SLS encompasses the NPH region of the core peptide. Interestingly, this region is found in all characterized TOMM cytolysins, as well as the novel putative TOMM cytolysins we discovered. We propose that this conserved region represents the defining feature of the SLS-like TOMM family. We demonstrate the cytolytic potential of a Bbsl SLS-like precursor peptide, which has a core region of similar length to the SLS minimal cytolytic unit, when modified with purified SLS biosynthetic enzymes. As such, we speculate that some Borrelia have the potential to produce a TOMM cytolysin, although the biological significance of this finding remains to be determined. In addition to providing new insight into the structure-activity relationships of SLS, this study greatly expands the cytolysin group of TOMMs.

No MeSH data available.


Related in: MedlinePlus