Limits...
Germline-encoded neutralization of a Staphylococcus aureus virulence factor by the human antibody repertoire

View Article: PubMed Central - PubMed

ABSTRACT

Staphylococcus aureus is both an important pathogen and a human commensal. To explore this ambivalent relationship between host and microbe, we analysed the memory humoral response against IsdB, a protein involved in iron acquisition, in four healthy donors. Here we show that in all donors a heavily biased use of two immunoglobulin heavy chain germlines generated high affinity (pM) antibodies that neutralize the two IsdB NEAT domains, IGHV4-39 for NEAT1 and IGHV1-69 for NEAT2. In contrast to the typical antibody/antigen interactions, the binding is primarily driven by the germline-encoded hydrophobic CDRH-2 motifs of IGHV1-69 and IGHV4-39, with a binding mechanism nearly identical for each antibody derived from different donors. Our results suggest that IGHV1-69 and IGHV4-39, while part of the adaptive immune system, may have evolved under selection pressure to encode a binding motif innately capable of recognizing and neutralizing a structurally conserved protein domain involved in pathogen iron acquisition.

No MeSH data available.


Related in: MedlinePlus

Characterization of anti-IsdB antibodies in the human memory B cell repertoire.(a) A persistent population of IsdB+ memory B cells from the peripheral blood mononuclear cells (PBMC) of a donor (D3) was observed over a 15-month period. By FACS, ∼0.06% of the IgM− CD19+ CD27+ memory B cells in the total memory B cell repertoire of this donor bind IsdB. (b) Most of the cloned BCR transcripts of the IsdB+ memory B cells collected at month 1, 3 and 15 are clonally related. BCR sequences from single-cell cloning of IsdB+ memory B cells were clustered based on heavy chain V-gene usage and CDR-H3 sequences. In total, we identified 31 unique clusters from donor D3 over the three collection time points. The Venn diagram shows that sibling clones within a cluster can be isolated at multiple time points. (c) Two distinct sets (bin C and bin P) of function-blocking antibodies specifically target NEAT1 and NEAT2, respectively. Single-cell cloning was performed at three different time points for donor D3 and one time each for donors D1, D2, and D4. In total, 75 unique antibodies targeting IsdB were identified and characterized. Shown here are the results of a comprehensive epitope binning analysis of 67 antibodies. Each reformatted clone is shown as a box and coloured according to its VH germline usage. The height of the box indicates the number of clustered BCR transcripts represented for each reformatted clone. There are in total 9, 34, 327, and 68 anti-IsdB single-cell BCR transcripts for D1, D2, D3 and D4, respectively. Each column of clones represents an epitope bin and this is overlaid on top of a linear representation of the IsdB molecule with NEAT1 in orange, and NEAT2 in blue. Clones that are able to fully block haemoglobin binding are outlined with a red box.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5120205&req=5

f1: Characterization of anti-IsdB antibodies in the human memory B cell repertoire.(a) A persistent population of IsdB+ memory B cells from the peripheral blood mononuclear cells (PBMC) of a donor (D3) was observed over a 15-month period. By FACS, ∼0.06% of the IgM− CD19+ CD27+ memory B cells in the total memory B cell repertoire of this donor bind IsdB. (b) Most of the cloned BCR transcripts of the IsdB+ memory B cells collected at month 1, 3 and 15 are clonally related. BCR sequences from single-cell cloning of IsdB+ memory B cells were clustered based on heavy chain V-gene usage and CDR-H3 sequences. In total, we identified 31 unique clusters from donor D3 over the three collection time points. The Venn diagram shows that sibling clones within a cluster can be isolated at multiple time points. (c) Two distinct sets (bin C and bin P) of function-blocking antibodies specifically target NEAT1 and NEAT2, respectively. Single-cell cloning was performed at three different time points for donor D3 and one time each for donors D1, D2, and D4. In total, 75 unique antibodies targeting IsdB were identified and characterized. Shown here are the results of a comprehensive epitope binning analysis of 67 antibodies. Each reformatted clone is shown as a box and coloured according to its VH germline usage. The height of the box indicates the number of clustered BCR transcripts represented for each reformatted clone. There are in total 9, 34, 327, and 68 anti-IsdB single-cell BCR transcripts for D1, D2, D3 and D4, respectively. Each column of clones represents an epitope bin and this is overlaid on top of a linear representation of the IsdB molecule with NEAT1 in orange, and NEAT2 in blue. Clones that are able to fully block haemoglobin binding are outlined with a red box.

Mentions: We first determined the presence of IsdB-reactive B cells in blood samples collected from a donor (D3) at months 1, 3 and 15 using flow cytometry (FACS), single-cell cloning (Supplementary Fig. 1) and high-throughput sequencing techniques. By FACS, we observed the persistence of a distinct IsdB-reactive memory B cell population (∼0.06%) within the IgM negative peripheral memory compartment (Fig. 1a). The majority of the IsdB-reactive memory B cells collected at three different time points expressed clonally related B cell receptor (BCR) transcripts: 25 of the 31 unique IsdB-reactive clusters obtained from this donor contained sibling transcripts isolated from at least two different time points (Fig. 1b). Longitudinal lineage analysis of the heavy chain variable region of these clusters indicates that the immune system maintains a repertoire of continually evolving antibodies against IsdB, presumably as a consequence of steady or intermittent exposure to low levels of antigen due to the commensal relationship between humans and S. aureus (Supplementary Fig. 2).


Germline-encoded neutralization of a Staphylococcus aureus virulence factor by the human antibody repertoire
Characterization of anti-IsdB antibodies in the human memory B cell repertoire.(a) A persistent population of IsdB+ memory B cells from the peripheral blood mononuclear cells (PBMC) of a donor (D3) was observed over a 15-month period. By FACS, ∼0.06% of the IgM− CD19+ CD27+ memory B cells in the total memory B cell repertoire of this donor bind IsdB. (b) Most of the cloned BCR transcripts of the IsdB+ memory B cells collected at month 1, 3 and 15 are clonally related. BCR sequences from single-cell cloning of IsdB+ memory B cells were clustered based on heavy chain V-gene usage and CDR-H3 sequences. In total, we identified 31 unique clusters from donor D3 over the three collection time points. The Venn diagram shows that sibling clones within a cluster can be isolated at multiple time points. (c) Two distinct sets (bin C and bin P) of function-blocking antibodies specifically target NEAT1 and NEAT2, respectively. Single-cell cloning was performed at three different time points for donor D3 and one time each for donors D1, D2, and D4. In total, 75 unique antibodies targeting IsdB were identified and characterized. Shown here are the results of a comprehensive epitope binning analysis of 67 antibodies. Each reformatted clone is shown as a box and coloured according to its VH germline usage. The height of the box indicates the number of clustered BCR transcripts represented for each reformatted clone. There are in total 9, 34, 327, and 68 anti-IsdB single-cell BCR transcripts for D1, D2, D3 and D4, respectively. Each column of clones represents an epitope bin and this is overlaid on top of a linear representation of the IsdB molecule with NEAT1 in orange, and NEAT2 in blue. Clones that are able to fully block haemoglobin binding are outlined with a red box.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5120205&req=5

f1: Characterization of anti-IsdB antibodies in the human memory B cell repertoire.(a) A persistent population of IsdB+ memory B cells from the peripheral blood mononuclear cells (PBMC) of a donor (D3) was observed over a 15-month period. By FACS, ∼0.06% of the IgM− CD19+ CD27+ memory B cells in the total memory B cell repertoire of this donor bind IsdB. (b) Most of the cloned BCR transcripts of the IsdB+ memory B cells collected at month 1, 3 and 15 are clonally related. BCR sequences from single-cell cloning of IsdB+ memory B cells were clustered based on heavy chain V-gene usage and CDR-H3 sequences. In total, we identified 31 unique clusters from donor D3 over the three collection time points. The Venn diagram shows that sibling clones within a cluster can be isolated at multiple time points. (c) Two distinct sets (bin C and bin P) of function-blocking antibodies specifically target NEAT1 and NEAT2, respectively. Single-cell cloning was performed at three different time points for donor D3 and one time each for donors D1, D2, and D4. In total, 75 unique antibodies targeting IsdB were identified and characterized. Shown here are the results of a comprehensive epitope binning analysis of 67 antibodies. Each reformatted clone is shown as a box and coloured according to its VH germline usage. The height of the box indicates the number of clustered BCR transcripts represented for each reformatted clone. There are in total 9, 34, 327, and 68 anti-IsdB single-cell BCR transcripts for D1, D2, D3 and D4, respectively. Each column of clones represents an epitope bin and this is overlaid on top of a linear representation of the IsdB molecule with NEAT1 in orange, and NEAT2 in blue. Clones that are able to fully block haemoglobin binding are outlined with a red box.
Mentions: We first determined the presence of IsdB-reactive B cells in blood samples collected from a donor (D3) at months 1, 3 and 15 using flow cytometry (FACS), single-cell cloning (Supplementary Fig. 1) and high-throughput sequencing techniques. By FACS, we observed the persistence of a distinct IsdB-reactive memory B cell population (∼0.06%) within the IgM negative peripheral memory compartment (Fig. 1a). The majority of the IsdB-reactive memory B cells collected at three different time points expressed clonally related B cell receptor (BCR) transcripts: 25 of the 31 unique IsdB-reactive clusters obtained from this donor contained sibling transcripts isolated from at least two different time points (Fig. 1b). Longitudinal lineage analysis of the heavy chain variable region of these clusters indicates that the immune system maintains a repertoire of continually evolving antibodies against IsdB, presumably as a consequence of steady or intermittent exposure to low levels of antigen due to the commensal relationship between humans and S. aureus (Supplementary Fig. 2).

View Article: PubMed Central - PubMed

ABSTRACT

Staphylococcus aureus is both an important pathogen and a human commensal. To explore this ambivalent relationship between host and microbe, we analysed the memory humoral response against IsdB, a protein involved in iron acquisition, in four healthy donors. Here we show that in all donors a heavily biased use of two immunoglobulin heavy chain germlines generated high affinity (pM) antibodies that neutralize the two IsdB NEAT domains, IGHV4-39 for NEAT1 and IGHV1-69 for NEAT2. In contrast to the typical antibody/antigen interactions, the binding is primarily driven by the germline-encoded hydrophobic CDRH-2 motifs of IGHV1-69 and IGHV4-39, with a binding mechanism nearly identical for each antibody derived from different donors. Our results suggest that IGHV1-69 and IGHV4-39, while part of the adaptive immune system, may have evolved under selection pressure to encode a binding motif innately capable of recognizing and neutralizing a structurally conserved protein domain involved in pathogen iron acquisition.

No MeSH data available.


Related in: MedlinePlus