scholarly journals Shared HIV envelope-specific B cell clonotypes induced by a pox-protein vaccine regimen

2021 ◽  
Author(s):  
Kristen W. Cohen ◽  
Lamar Ballweber-Fleming ◽  
Michael Duff ◽  
Rachael E. Whaley ◽  
Aaron Seese ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Neutralizing Antibodies ◽  
Rational Design ◽  
Critical Role ◽  
Memory B Cells ◽  
Protein Vaccine ◽  
Cell Repertoire ◽  
B Cell Repertoire ◽  
Specific Igg

An effective HIV-1 vaccine will likely induce potent, broad neutralizing antibodies. No candidate vaccines have elicited these responses presumably because they fail to activate human B cell precursors that can affinity mature to generate broad neutralizing antibodies. To identify the B cell clonotypes that are elicited, we conducted in-depth analyses of the envelope-specific B cell repertoire in recipients of ALVAC-HIV vector (vCP2438) and bivalent subtype C gp120 protein (HVTN100). We observed high frequencies of envelope-specific IgG+ memory B cells with restricted immunogenetic diversity, relative to non-vaccine induced memory B cells, with preferential expansions of distinct variable genes but limited accumulation of mutations. Many envelope-specific clonotypes were shared across vaccinees, but did not overlap with the envelope-negative memory repertoire, within and across subjects. Single-cell sequencing of envelope-specific IgG+ memory B cells often revealed VH1-2*02 and VK3-20 sequence co-expression and in one case, contained a 5 amino acid CDRL3, the canonical signature of VRC01-class antibodies, confirming that these B cells are extremely rare but detectable. Our study provides evidence that immunogens play a critical role in selecting and restricting the responding B cell repertoire and supports the rational design of HIV vaccines targeting specific B cell lineages for induction of broadly-reactive neutralizing antibodies.

scholarly journals Broad sarbecovirus neutralizing antibodies define a key site of vulnerability on the SARS-CoV-2 spike protein

2020 ◽  
Author(s):  
Anna Z. Wec ◽  
Daniel Wrapp ◽  
Andrew S. Herbert ◽  
Daniel Maurer ◽  
Denise Haslwanter ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Rational Design ◽  
Somatic Hypermutation ◽  
Protein A ◽  
Neutralizing Antibody ◽  
Memory B Cells ◽  
Antibody Responses ◽  
Cell Repertoire ◽  
Human Coronavirus ◽  
B Cell Repertoire

Broadly protective vaccines against known and pre-emergent coronaviruses are urgently needed. Critical to their development is a deeper understanding of cross-neutralizing antibody responses induced by natural human coronavirus (HCoV) infections. Here, we mined the memory B cell repertoire of a convalescent SARS donor and identified 200 SARS-CoV-2 binding antibodies that target multiple conserved sites on the spike (S) protein. A large proportion of the antibodies display high levels of somatic hypermutation and cross-react with circulating HCoVs, suggesting recall of pre-existing memory B cells (MBCs) elicited by prior HCoV infections. Several antibodies potently cross-neutralize SARS-CoV, SARS-CoV-2, and the bat SARS-like virus WIV1 by blocking receptor attachment and inducing S1 shedding. These antibodies represent promising candidates for therapeutic intervention and reveal a new target for the rational design of pan-sarbecovirus vaccines.

Profiling the Autoantibody Repertoire of Acquired Thrombotic Thrombocytopenic Purpura (aTTP) Patients By Single Cell Sorting and Deep Sequencing of Splenic B-Cells

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 108-108
Author(s):  
Magdalena Skowronska ◽  
Monica M Schaller ◽  
Johanna A. Kremer Hovinga Strebel
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Population ◽  
Memory B Cells ◽  
Light Chains ◽  
Cell Populations ◽  
Cell Repertoire ◽  
Specific Memory ◽  
B Cell Repertoire ◽  
Single Antigen

Abstract Introduction: Antibodies are the primary effector molecules in the humoral immune system. To create a diversity of receptors-antibodies B-cells undergo V(D)J gene recombination and somatic hypermutation. This allows the recognition of most of pathogens but also sometimes of self-antigen, which can lead to autoimmune diseases. Autoantibodies (Abs) neutralizing and/or accelerating the clearance of ADAMTS13 are present in nearly all acquired thrombotic thrombocytopenic purpura (aTTP) patients with severe ADAMTS13 activity (<5%). As increasing evidence points at the spleen as a major reservoir for antigen specific memory B-cells, we investigated the splenic B-cell repertoire. We sequenced heavy and corresponding light chains from single antigen-specific memory B-cells and deep sequenced global antibody repertoire among different B-cell populations. Combining both, the single antigen-specific repertoire and the general global repertoire from the same patients specimens allows an investigation of the V(D)J gene usage and abundance of specific V(D)J junction in the B-cell populations within one patient and also among different patients. Analysis of the repertoire enables better understanding of the humoral autoimmune response. Methods: Specimens were analyzed from 8 aTTP patients who were splenectomized due to a refractory course of the autoimmune disease. Splenic mononuclear cells were sorted by flow-cytometry into naïve and transitional (CD27-, IgD+), unswitched memory (CD27+, IgD+) and switched memory (CD27+, IgD-) as well as (CD27+, CD20-/CD19-, CD38+) plasma cells. The frequencies of highly positive anti-ADAMTS13 B-cells in each B-cell population were calculated. Library consisting of antibody cDNA from each B-cell population was deep sequenced on MiSeq Ilumina. Prime analysis were performed in CLC Genomics Workbench. High quality sequences were submitted to IMGT/high V-QUEST web-based analysis tool to determine for V(D)J genes alignments, and V(D)J junction (antigen binding region) and mutations analysis. The IMGT output was parsed into MySQL database for further analysis. In addition, from 4 patients anti-ADAMTS13 specific IgG bearing cells were individually sorted. Gene transcripts of single cells were reverse-transcribed followed by nested PCR of IgG heavy/light chains. V(D)J pairings were visualized with Circos software. In MySQL we compared single cell antigen specific sequence with global repertoire. Results: Anti-ADAMTS13 specific B-cells were detected in the spleen of all patients (average 0.01% of total B-cell population). Deep sequencing of the total B-cell repertoire revealed ~3 million productive sequences, from which ~2 million were unique. Splenic anti-ADAMTS13 specific B-cells of four aTTP patients revealed 80 antibodies with unique V(D)J junction. Among those most frequently used V-genes were IGHV1-69 and IGHV3-30 (15% and 12% respectively) in global repertoire 1.5% of antibodies were encoded with IGHV1-69 gene within respective population. The average identity to germline was lower in ADMTST13 specific B-cells (92% compare to 96%). Four V(D)J junctions were convergent in at least 2 patients (identical amino acid sequence). Conclusion: Anti-ADAMTS13 specific B-cells were found in all aTTP patients in different B-cell populations. We observed enrichment of some variable gene segments when comparing the specific anti-ADAMTS13 to the total splenic repertoire (mainly IGHV1-69). Finding convergent V(D)J junction is very promising and also confirms previous finding of our group where similar V(D)J junction were found among two patients. Currently we clone selected single sorted monoclonal Abs (Schaller et al, Blood 2014;124(23):3469-79). Functional testing will allow the selection of the inhibitory Abs to be used as tools to develop anti-idiotypic specific therapies for aTTP patients. Disclosures No relevant conflicts of interest to declare.

scholarly journals A public antibody class recognizes a novel S2 epitope exposed on open conformations of SARS-CoV-2 spike

2021 ◽  
Author(s):  
Mathieu Claireaux ◽  
Tom G Caniels ◽  
Marlon de Gast ◽  
Julianna Han ◽  
Denise Guerra ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Receptor ◽  
Memory B Cells ◽  
B Cell Receptor ◽  
Cell Repertoire ◽  
Fold Enrichment ◽  
B Cell Repertoire ◽  
High Fraction ◽  
Antibody Class

AbstractDelineating the origins and properties of antibodies elicited by SARS-CoV-2 infection and vaccination is critical for understanding their benefits and potential shortcomings. Therefore, we investigated the SARS-CoV-2 spike (S)-reactive B cell repertoire in unexposed individuals by flow cytometry and single-cell sequencing. We found that ∼82% of SARS-CoV-2 S-reactive B cells show a naive phenotype, which represents an unusually high fraction of total human naive B cells (∼0.1%). Approximately 10% of these naive S-reactive B cells shared an IGHV1-69/IGKV3-11 B cell receptor pairing, an enrichment of 18-fold compared to the complete naive repertoire. A proportion of memory B cells, comprising switched (∼0.05%) and unswitched B cells (∼0.04%), was also reactive with S and some of these cells were reactive with ADAMTS13, which is associated with thrombotic thrombocytopenia. Following SARS-CoV-2 infection, we report an average 37-fold enrichment of IGHV1-69/IGKV3-11 B cell receptor pairing in the S-reactive memory B cells compared to the unselected memory repertoire. This class of B cells targets a previously undefined non-neutralizing epitope on the S2 subunit that becomes exposed on S proteins used in approved vaccines when they transition away from the native pre-fusion state because of instability. These findings can help guide the improvement of SARS-CoV-2 vaccines.

scholarly journals Efficient Peripheral Clonal Elimination of B Lymphocytes in MRL/lpr Mice Bearing Autoantibody Transgenes

1998 ◽  
Vol 188 (5) ◽  
pp. 909-917 ◽  
Author(s):  
Jennifer A. Kench ◽  
David M. Russell ◽  
David Nemazee
Keyword(s):  
B Cells ◽  
B Cell ◽  
Genetic Background ◽  
Cell Tolerance ◽  
Cell Repertoire ◽  
B Cell Tolerance ◽  
B Cell Repertoire ◽  
Nuclear Antigens ◽  
Large Numbers ◽  
Liver And Kidney

Peripheral B cell tolerance was studied in mice of the autoimmune-prone, Fas-deficient MRL/ lpr.H-2d genetic background by introducing a transgene that directs expression of membrane-bound H-2Kb antigen to liver and kidney (MT-Kb) and a second transgene encoding antibody reactive with this antigen (3-83μδ, anti-Kk,b). Control immunoglobulin transgenic (Ig-Tg) MRL/lpr.H-2d mice lacking the Kb antigen had large numbers of splenic and lymph node B cells bearing the transgene-encoded specificity, whereas B cells of the double transgenic (Dbl-Tg) MRL/lpr.H-2d mice were deleted as efficiently as in Dbl-Tg mice of a nonautoimmune B10.D2 genetic background. In spite of the severely restricted peripheral B cell repertoire of the Ig-Tg MRL/lpr.H-2d mice, and notwithstanding deletion of the autospecific B cell population in the Dbl-Tg MRL/lpr.H-2d mice, both types of mice developed lymphoproliferation and exhibited elevated levels of IgG anti-chromatin autoantibodies. Interestingly, Dbl-Tg MRL/lpr.H-2d mice had a shorter lifespan than Ig-Tg MRL/lpr.H-2d mice, apparently as an indirect result of their relative B cell lymphopenia. These data suggest that in MRL/lpr mice peripheral B cell tolerance is not globally defective, but that certain B cells with receptors specific for nuclear antigens are regulated differently than are cells reactive to membrane autoantigens.

scholarly journals Memory B Cell Responses to Vibrio cholerae O1 Lipopolysaccharide Are Associated with Protection against Infection from Household Contacts of Patients with Cholera in Bangladesh

2012 ◽  
Vol 19 (6) ◽  
pp. 842-848 ◽  
Author(s):  
Sweta M. Patel ◽  
Mohammad Arif Rahman ◽  
M. Mohasin ◽  
M. Asrafuzzaman Riyadh ◽  
Daniel T. Leung ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Vibrio Cholerae ◽  
Memory B Cells ◽  
Content Type ◽  
Cell Responses ◽  
Household Contacts ◽  
Specific Memory ◽  
Specific Igg ◽  
Protection Against Infection

ABSTRACTVibrio choleraeO1 causes cholera, a dehydrating diarrheal disease. We have previously shown thatV. cholerae-specific memory B cell responses develop after cholera infection, and we hypothesize that these mediate long-term protective immunity against cholera. We prospectively followed household contacts of cholera patients to determine whether the presence of circulatingV. choleraeO1 antigen-specific memory B cells on enrollment was associated with protection againstV. choleraeinfection over a 30-day period. Two hundred thirty-six household contacts of 122 index patients with cholera were enrolled. The presence of lipopolysaccharide (LPS)-specific IgG memory B cells in peripheral blood on study entry was associated with a 68% decrease in the risk of infection in household contacts (P= 0.032). No protection was associated with cholera toxin B subunit (CtxB)-specific memory B cells or IgA memory B cells specific to LPS. These results suggest that LPS-specific IgG memory B cells may be important in protection against infection withV. choleraeO1.

scholarly journals Mouse APOBEC3 affects the production of virus-neutralizing antibodies by restricting early retroviral replication, not by altering the B-cell repertoire

Retrovirology ◽  
2009 ◽  
Vol 6 (S2) ◽  
Author(s):  
Masaaki Miyazawa ◽  
Sachiyo Tsuji-Kawahara ◽  
Tomomi Chikaishi ◽  
Maiko Kato ◽  
Shiki Takamura
Keyword(s):  
B Cell ◽  
Neutralizing Antibodies ◽  
Cell Repertoire ◽  
B Cell Repertoire ◽  
Retroviral Replication

scholarly journals High-frequency representation of a single VH gene in the expressed human B cell repertoire.

1993 ◽  
Vol 177 (2) ◽  
pp. 409-418 ◽  
Author(s):  
A K Stewart ◽  
C Huang ◽  
B D Stollar ◽  
R S Schwartz
Keyword(s):  
B Cells ◽  
B Cell ◽  
High Frequency ◽  
Gene Segment ◽  
Normal Subjects ◽  
Cell Repertoire ◽  
B Cell Repertoire ◽  
V Genes ◽  
Vh Gene ◽  
Vh Genes

Idiotype (Id) 16/6 marks a variable (V) region structure that occurs frequently in the human immunoglobulin repertoire. The basis of the Id has been traced to a germline heavy chain gene segment, VH18/2 (VH26). To pursue the molecular basis for the frequency of Id 16/6, we have analyzed polymerase chain reaction-generated C mu, C gamma, and VH3 family V gene libraries derived from the circulating and tonsillar B cells of four normal individuals and from the B cells of two patients with active systemic lupus erythematosus (SLE). The frequency of VH18/2 in these libraries was compared with three control VH genes, VH56P1, VH21/28, and VHA57. Plaque lifts from C mu and C gamma VH cDNA libraries were screened with gene-specific oligonucleotide probes. The frequency of VH18/2 ranged from 4 to 10% of JH+ plaques (two of five times that of control VH genes). In four VH3 family-specific libraries derived from rearranged DNA, VH18/2 represented 19-33% of VH3+ plaques. Hybridizing VH18/2 plaques were 98-100% homologous to the germline VH gene; mutations when present were often in framework 3. Extensive variation was seen in the complementarity determining region 3 sequences of these rearranged V genes. The high frequency of VH18/2 expression in the B cell repertoire was confirmed by sequencing randomly picked JH+ plaques. In two patients with active SLE the frequency of use of VH18/2 was not greater than that observed in normal subjects. These results show that VH18/2 is overrepresented in the B cell repertoire of normal subjects and suggest that the immune repertoire may be dominated by relatively few V genes.

scholarly journals SARS-CoV-2 spike-specific memory B cells express higher levels of T-bet and FcRL5 after non-severe COVID-19 as compared to severe disease

PLoS ONE ◽  
2021 ◽  
Vol 16 (12) ◽  
pp. e0261656
Author(s):  
Raphael A. Reyes ◽  
Kathleen Clarke ◽  
S. Jake Gonzales ◽  
Angelene M. Cantwell ◽  
Rolando Garza ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Symptom Onset ◽  
Cell Response ◽  
Severe Disease ◽  
Memory B Cells ◽  
Memory B Cell ◽  
Specific Memory ◽  
Specific Igg ◽  
B Cell Response

SARS-CoV-2 infection elicits a robust B cell response, resulting in the generation of long-lived plasma cells and memory B cells. Here, we aimed to determine the effect of COVID-19 severity on the memory B cell response and characterize changes in the memory B cell compartment between recovery and five months post-symptom onset. Using high-parameter spectral flow cytometry, we analyzed the phenotype of memory B cells with reactivity against the SARS-CoV-2 spike protein or the spike receptor binding domain (RBD) in recovered individuals who had been hospitalized with non-severe (n = 8) or severe (n = 5) COVID-19. One month after symptom onset, a substantial proportion of spike-specific IgG+ B cells showed an activated phenotype. In individuals who experienced non-severe disease, spike-specific IgG+ B cells showed increased expression of markers associated with durable B cell memory, including T-bet and FcRL5, as compared to individuals who experienced severe disease. While the frequency of T-bet+ spike-specific IgG+ B cells differed between the two groups, these cells predominantly showed an activated switched memory B cell phenotype in both groups. Five months post-symptom onset, the majority of spike-specific memory B cells had a resting phenotype and the percentage of spike-specific T-bet+ IgG+ memory B cells decreased to baseline levels. Collectively, our results highlight subtle differences in the B cells response after non-severe and severe COVID-19 and suggest that the memory B cell response elicited during non-severe COVID-19 may be of higher quality than the response after severe disease.

scholarly journals The pre-existing human antibody repertoire to computationally optimized influenza H1 hemagglutinin vaccines

2021 ◽  
Author(s):  
Kaito Nagashima ◽  
John V Dzimianski ◽  
Julianna Han ◽  
Nada Abbadi ◽  
Aaron D Gingerich ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cross Reactivity ◽  
Influenza Viruses ◽  
Human Antibody ◽  
Protein Vaccine ◽  
Cell Repertoire ◽  
Neutralizing Activity ◽  
Human B Cells ◽  
Repertoire Sequencing

The computationally optimized broadly reactive antigen (COBRA) approach has previously been used to generate hemagglutinin (HA) immunogens for several influenza subtypes that expand vaccine-elicited antibody breadth. As nearly all individuals have pre-existing immunity to influenza viruses, influenza-specific memory B cells will likely be recalled upon COBRA HA vaccination. We determined the epitope specificity and repertoire characteristics of pre-existing human B cells to H1 COBRA HA antigens. Cross-reactivity between wild type HA and H1 COBRA HA proteins were observed at both the oligoclonal B cell level and for a subset of isolated monoclonal antibodies (mAbs). The mAbs bound five distinct epitopes on the pandemic A/California/04/2009 head and stem domains, and the majority of the mAbs had HAI and neutralizing activity against pandemic H1 strains. Two head-directed mAbs, CA09-26 and CA09-45, had HAI and neutralizing activity against a pre-pandemic H1 strain. One mAb, P1-05, targets the stem region of H1 HA proteins, but does not compete with known stem-targeting H1 mAbs. We determined that mAb P1-05 recognizes a recently discovered membrane proximal epitope on HA, the anchor epitope, and we identified similar mAbs using B cell repertoire sequencing. In addition, the trimerization domain distance from HA was critical to recognition of this epitope by P1-05. Overall, these data indicate that seasonally vaccinated individuals possess a population of functional H1 COBRA HA-reactive B cells that target head, central stalk, and anchor epitopes, and demonstrate the importance of structure-based assessment of subunit protein vaccine candidates to ensure accessibility of optimal protein epitopes.

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