Biology subjects

Csepregi, L.

Publications and source records attributed to Csepregi, L..

6 recordsLinked to original sources

bioRxiv2021

The Physiological Landscape and Specificity of Antibody Repertoires

Diverse antibody repertoires spanning multiple lymphoid organs (e.g., bone marrow, spleen, lymph nodes) form the foundation of protective humoral immunity. Changes in their composition across lymphoid organs are a consequence of B-cell selection and migration events leading to a highly dynamic and unique physiological landscape of antibody repertoires upon antigenic challenge (e.g., vaccination). However, to what extent B cells encoding identical or similar antibody sequences (clones) are distributed across multiple lymphoid organs and how this is shaped by the strength of a humoral response, remains largely unexplored. Here, we performed an in-depth systems analysis of antibody repertoires across multiple distinct lymphoid organs of immunized mice, and discovered that organ-specific antibody repertoire features (e.g., germline V-gene usage and clonal expansion profiles) equilibrated upon a strong humoral response (multiple immunizations and high serum titers). This resulted in a surprisingly high degree of repertoire consolidation, characterized by highly connected and overlapping B-cell clones across multiple lymphoid organs. Finally, we revealed distinct physiological axes indicating clonal migrations and showed that antibody repertoire consolidation directly correlated with antigen-specificity. Our study uncovered how a strong humoral response resulted in a more uniform but redundant physiological landscape of antibody repertoires, indicating that increases in antibody serum titers were a result of synergistic contributions from antigen-specific B-cell clones distributed across multiple lymphoid organs. Our findings provide valuable insights for the assessment and design of vaccine strategies.

immunology↗

bioRxiv2021

Phenotypic determinism and stochasticity in antibody repertoires of clonally expanded plasma cells

The capacity of humoral B cell-mediated immunity to effectively respond to and protect against pathogenic infections is largely driven by the presence of a diverse repertoire of polyclonal antibodies in the serum, which are produced by plasma cells (PCs).1,2 Recent studies have started to reveal the balance between deterministic mechanisms and stochasticity of antibody repertoires on a genotypic level (i.e., clonal diversity, somatic hypermutation, germline gene usage).3-8 However, it remains unclear if clonal selection and expansion of PCs follows any deterministic rules or is stochastic with regards to phenotypic antibody properties (i.e., antigen-binding, affinity, epitope specificity). Here we report on the in-depth genotypic and phenotypic characterization of clonally expanded PC antibody repertoires following protein immunization. We find that there is only a strong correlation with antigen-specificity among the most expanded clones (top ~ 10), whereas among the rest of the clonal repertoire antigen-specificity is stochastic. Furthermore, we report both on a polyclonal repertoire and clonal lineage level that antibody-antigen binding affinity does not correlate with clonal expansion or somatic hypermutation. Lastly, we provide evidence for convergence towards dominant epitopes despite clonal sequence diversity among the most expanded clones. Our results highlight the extent to which clonal expansion can be ascribed to antigen binding, affinity and epitope specificity and they have implications for the assessment of effective vaccines. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=199 SRC="FIGDIR/small/452687v1_ufig1.gif" ALT="Figure 1"> View larger version (57K): org.highwire.dtl.DTLVardef@e907c1org.highwire.dtl.DTLVardef@fc45b6org.highwire.dtl.DTLVardef@44bd91org.highwire.dtl.DTLVardef@f98f3e_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2021

Single-cell sequencing reveals clonally expanded plasma cells during chronic viral infection produce virus-specific and cross-reactive antibodies

Plasma cells and their secreted antibodies play a central role in the long-term protection against chronic viral infection. However, due to experimental limitations, a comprehensive description of linked genotypic, phenotypic, and antibody repertoire features of plasma cells (gene expression, clonal frequency, virus specificity, and affinity) has been challenging to obtain. To address this, we performed single-cell transcriptome and antibody repertoire sequencing of the murine bone marrow plasma cell population following chronic lymphocytic choriomeningitis virus infection. Our single-cell sequencing approach recovered full-length and paired heavy and light chain sequence information for thousands of plasma cells and enabled us to perform recombinant antibody expression and specificity screening. Antibody repertoire analysis revealed that, relative to protein immunization, chronic infection led to increased levels of clonal expansion, class-switching, and somatic variants. Furthermore, antibodies from the highly expanded and class-switched (IgG) plasma cells were found to be specific for multiple viral antigens and a subset of clones exhibited cross-reactivity to non-viral- and auto-antigens. Integrating single-cell transcriptome data with antibody specificity suggested that plasma cell transcriptional phenotype was correlated to viral antigen specificity. Our findings demonstrate that chronic viral infection can induce and sustain plasma cell clonal expansion, combined with significant somatic hypermutation, and can generate cross-reactive antibodies. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=69 SRC="FIGDIR/small/428852v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@9d0852org.highwire.dtl.DTLVardef@18d46fcorg.highwire.dtl.DTLVardef@2f7f80org.highwire.dtl.DTLVardef@5be3fd_HPS_FORMAT_FIGEXP M_FIG Graphical abstract.Single-cell sequencing reveals clonally expanded plasma cells during chronic viral infection produce virus-specific and cross-reactive antibodies. C_FIG

immunology↗

bioRxiv2020

Convergent selection in antibody repertoires is revealed by deep learning

Adaptive immunity is driven by the ability of lymphocytes to undergo V(D)J recombination and generate a highly diverse set of immune receptors (B cell receptors/secreted antibodies and T cell receptors) and their subsequent clonal selection and expansion upon molecular recognition of foreign antigens. These principles lead to remarkable, unique and dynamic immune receptor repertoires1. Deep sequencing provides increasing evidence for the presence of commonly shared (convergent) receptors across individual organisms within one species2-4. Convergent selection of specific receptors towards various antigens offers one explanation for these findings. For example, single cases of convergence have been reported in antibody repertoires of viral infection or allergy5-8. Recent studies demonstrate that convergent selection of sequence motifs within T cell receptor (TCR) repertoires can be identified on an even wider scale9,10. Here we report that there is extensive convergent selection in antibody repertoires of mice for a range of protein antigens and immunization conditions. We employed a deep learning approach utilizing variational autoencoders (VAEs) to model the underlying process of B cell receptor (BCR) recombination and assume that the data generation follows a Gaussian mixture model (GMM) in latent space. This provides both a latent embedding and cluster labels that group similar sequences, thus enabling the discovery of a multitude of convergent, antigen-associated sequence patterns. Using a linear, one-versus-all support vector machine (SVM), we confirm that the identified sequence patterns are predictive of antigenic exposure and outperform predictions based on the occurrence of public clones. Recombinant expression of both natural and in silico-generated antibodies possessing convergent patterns confirms their binding specificity to target antigens. Our work highlights to which extent convergence in antibody repertoires can occur and shows how deep learning can be applied for immunodiagnostics and antibody discovery and engineering.

immunology↗

bioRxiv2019

Molecular design, optimization and genomic integration of chimeric B cell receptors in murine B cells

Immune cell therapies based on the integration of synthetic antigen receptors provide a powerful strategy for the treatment of diverse diseases, most notably retargeting T cells engineered to express chimeric antigen receptors (CAR) for cancer therapy. In addition to T lymphocytes, B lymphocytes may also represent valuable immune cells that can be engineered for therapeutic purposes such as protein replacement therapy or recombinant antibody production. In this article, we report a promising concept for the molecular design, optimization and genomic integration of a novel class of synthetic antigen receptors, chimeric B cell receptors (CBCR). We initially optimized CBCR expression and detection by modifying the extracellular surface tag, the transmembrane regions and intracellular signaling domains. For this purpose, we stably integrated a series of CBCR variants into immortalized B cell hybridomas using CRISPR-Cas9. Subsequently, we developed a reliable and consistent pipeline to precisely introduce cassettes of several kilobases size into the genome of primary murine B cells, again via CRISPR-Cas9 induced HDR. Finally, we were able to show the robust surface expression and antigen recognition of a synthetic CBCR in primary B cells. We anticipate that CBCRs and our approach for engineering primary B cells will be a valuable tool for the advancement of future B cell-based immune therapies.

immunology↗

bioRxiv2018

Boosting subdominant neutralizing antibody responses with a computationally designed epitope-focused immunogen

Throughout the last decades, vaccination has been key to prevent and eradicate infectious diseases. However, many pathogens (e.g. respiratory syncytial virus (RSV), influenza, dengue and others) have resisted vaccine development efforts, largely due to the failure to induce potent antibody responses targeting conserved epitopes. Deep profiling of human B-cells often reveals potent neutralizing antibodies that emerge from natural infection, but these specificities are generally subdominant (i.e., are present in low titers). A major challenge for next-generation vaccines is to overcome established immunodominance hierarchies and focus antibody responses on crucial neutralization epitopes. Here, we show that a computationally designed epitope-focused immunogen presenting a single RSV neutralization epitope elicits superior epitope-specific responses compared to the viral fusion protein. In addition, the epitope-focused immunogen efficiently boosts antibodies targeting the Palivizumab epitope, resulting in enhanced neutralization. Overall, we show that epitope-focused immunogens can boost subdominant neutralizing antibody responses in vivo and reshape established antibody hierarchies.

immunology↗