B cell–derived IL-27 promotes control of persistent LCMV infection

Recent studies have identified a critical role for B cell–produced cytokines in regulating both humoral and cellular immunity. Here, we show that B cells are an essential source of interleukin-27 (IL-27) during persistent lymphocytic choriomeningitis virus (LCMV) clone 13 (Cl-13) infection. By using conditional knockout mouse models with specific IL-27p28 deletion in B cells, we observed that B cell–derived IL-27 promotes survival of virus-specific CD4 T cells and supports functions of T follicular helper (Tfh) cells. Mechanistically, B cell–derived IL-27 promotes CD4 T cell function, antibody class switch, and the ability to control persistent LCMV infection. Deletion of IL-27ra in T cells demonstrated that T cell–intrinsic IL-27R signaling is essential for viral control, optimal CD4 T cell responses, and antibody class switch during persistent LCMV infection. Collectively, our findings identify a cellular mechanism whereby B cell–derived IL-27 drives antiviral immunity and antibody responses through IL-27 signaling on T cells to promote control of LCMV Cl-13 infection.

Isolation and comparative analysis of antibodies that broadly neutralize sarbecoviruses

The devastation caused by SARS-CoV-2 has made clear the importance of pandemic preparedness. To address future zoonotic outbreaks due to related viruses in the sarbecovirus subgenus, we identified a human monoclonal antibody, 10-40, that neutralized or bound all sarbecoviruses tested in vitro and protected against SARS-CoV-2 and SARS-CoV in vivo. Comparative studies with other receptor-binding domain (RBD)-directed antibodies showed 10-40 to have the greatest breadth against sarbecoviruses and thus its promise as an agent for pandemic preparedness. Moreover, structural analyses on 10-40 and similar antibodies not only defined an epitope cluster in the inner face of the RBD that is well conserved among sarbecoviruses, but also uncovered a new antibody class with a common CDRH3 motif. Our analyses also suggested that elicitation of this class of antibodies may not be overly difficult, an observation that bodes well for the development of a pan-sarbecovirus vaccine.

Characterization of the significant decline in humoral immune response six months post-SARS-CoV-2 mRNA vaccination: A systematic review

Accumulating evidence shows a progressive decline in the efficacy of coronavirus disease 2019 (COVID-19) mRNA vaccines such as Pfizer-BioNTech (mRNA BNT161b2) and Moderna (mRNA-1273) in preventing breakthrough infections due to diminishing humoral immunity over time. Thus, this review characterizes the kinetics of anti-SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2) antibodies after the second dose of a primary cycle of COVID-19 mRNA vaccination. A systematic search of literature was performed and a total of 18 studies (N=15,980) were identified and reviewed. The percent difference of means of reported antibody titers were then calculated to determine the decline in humoral response after the peak levels post-vaccination. Findings revealed that the peak humoral response was reached at 21-28 days after the second dose, after which serum levels progressively diminished at 4-6 months post-vaccination. Additionally, results showed that regardless of age, sex, serostatus and presence of comorbidities, longitudinal data reporting antibody measurement exhibited a decline of both anti-receptor binding domain (RBD) IgG and anti-spike IgG, ranging from 94-95% at 90-180 days and 55-85% at 140-160 days, respectively, after the peak antibody response. This suggests that the rate of antibody decline may be independent of patient-related factors and peak antibody titers but mainly a function of time and antibody class/molecular target. Hence, this study highlights the necessity of more efficient vaccination strategies to provide booster administration in attenuating the effects of waning immunity, especially in the appearance of new variants of concerns (VoCs).

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

Delineating 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.

Multiple DSB Resection Activities Redundantly Promote Alternative End Joining-Mediated Class Switch Recombination

Alternative end joining (A-EJ) catalyzes substantial level of antibody class switch recombination (CSR) in B cells deficient for classical non-homologous end joining, featuring increased switch (S) region DSB resection and junctional microhomology (MH). While resection has been suggested to initiate A-EJ in model DSB repair systems using engineered endonucleases, the contribution of resection factors to A-EJ-mediated CSR remains unclear. In this study, we systematically dissected the requirement for individual DSB resection factors in A-EJ-mediated class switching with a cell-based assay system and high-throughput sequencing. We show that while CtIP and Mre11 both are mildly required for CSR in WT cells, they play more critical roles in mediating A-EJ CSR, which depend on the exonuclease activity of Mre11. While DNA2 and the helicase/HRDC domain of BLM are required for A-EJ by mediating long S region DSB resection, in contrast, Exo1’s resection-related function does not play any obvious roles for class switching in either c-NHEJ or A-EJ cells, or mediated in an AID-independent manner by joining of Cas9 breaks. Furthermore, ATM and its kinase activity functions at least in part independent of CtIP/Mre11 to mediate A-EJ switching in Lig4-deficient cells. In stark contrast to Lig4 deficiency, 53BP1-deficient cells do not depend on ATM/Mre11/CtIP for residual joining. We discuss the roles for each resection factor in A-EJ-mediated CSR and suggest that the extent of requirements for resection is context dependent.

Molecular strategies for antibody binding and escape of SARS-CoV-2 and its mutations

The COVID19 pandemic, caused by SARS-CoV-2, has infected more than 200 million people worldwide. Due to the rapid spreading of SARS-CoV-2 and its impact, it is paramount to find effective treatments against it. Human neutralizing antibodies are an effective method to fight viral infection. However, the recent discovery of new strains that substantially change the S-protein sequence has raised concern about vaccines and antibodies’ effectiveness. Here, using molecular simulations, we investigated the binding mechanisms between the S-protein and several antibodies. Multiple mutations were included to understand the strategies for antibody escape in new variants. We found that the combination of mutations K417N, E484K, L452R, and T478K produced higher binding energy to ACE2 than the wild type, suggesting higher efficiency to enter host cells. The mutations’ effect depends on the antibody class. While Class I enhances the binding avidity in the presence of N501Y mutation, class II antibodies showed a sharp decline in the binding affinity. Our simulations suggest that Class I antibodies will remain effective against the new strains. In contrast, Class II antibodies will have less affinity to the S-protein, potentially affecting these antibodies’ efficiency.

SARS-CoV-2 Beta variant infection elicits potent lineage-specific and cross-reactive antibodies

SARS-CoV-2 Beta variant of concern (VOC) resists neutralization by major classes of antibodies from non-VOC COVID-19 patients and vaccinated individuals. Here, serum of Beta variant infected patients revealed reduced cross-neutralization of non-VOC virus. From these patients, we isolated Beta-specific and cross-reactive receptor-binding domain (RBD) antibodies. The Beta-specificity results from recruitment of novel VOC-specific clonotypes and accommodation of VOC-defining amino acids into a major non-VOC antibody class that is normally sensitive to these mutations. The Beta-elicited cross-reactive antibodies share genetic and structural features with non-VOC-elicited antibodies, including a public VH1-58 clonotype targeting the RBD ridge independent of VOC mutations. These findings advance our understanding of the antibody response to SARS-CoV-2 shaped by antigenic drift with implications for design of next-generation vaccines and therapeutics.

The ASCIZ-DYNLL1 axis is essential for TLR4-mediated antibody responses and NF-κB pathway activation

Toll-like receptors (TLRs) and IL-1 receptors regulate immune and inflammatory responses by activating the nuclear factor (NF)-κB pathway. Here we report that B cell-specific loss of Dynein light chain-1 (DYNLL1, LC8) or its designated transcription factor ASCIZ (ATMIN) leads to severely reduced in vivo antibody responses to TLR4-dependent but not T cell-dependent antigens in mice. This defect was independent of DYNLL1’s established roles in modulating BIM-dependent apoptosis and 53BP1-dependent antibody class-switch recombination. In B cells and fibroblasts, the ASCIZ-DYNLL1 axis was required for TLR4-, IL-1- and CD40-mediated NF-κB pathway activation but dispensable for antigen receptor and TNF-α signalling. In contrast to previous reports that overexpressed DYNLL1 directly inhibits the phosphorylation and degradation of the NF-κB inhibitor IκBα, we found here that - under physiological conditions - DYNLL1 is required for signal-specific activation of the NF-κB pathway upstream of IκBα. Our data identify DYNLL1 as a signal-specific regulator of the NF-κB pathway and indicate that it may act as a universal modulator of TLR4 (and IL-1) signalling with wide-ranging roles in inflammation and immunity.

Antibody Class(es) Predictor for Epitopes (AbCPE): A Multi-Label Classification Algorithm

Development of vaccines and therapeutic antibodies to deal with infectious and other diseases are the most perceptible scientific interventions that have had huge impact on public health including that in the current Covid-19 pandemic. From inactivation methodologies to reverse vaccinology, vaccine development strategies of 21st century have undergone several transformations and are moving towards rational design approaches. These developments are driven by data as the combinatorials involved in antigenic diversity of pathogens and immune repertoire of hosts are enormous. The computational prediction of epitopes is central to these developments and numerous B-cell epitope prediction methods developed over the years in the field of immunoinformatics have contributed enormously. Most of these methods predict epitopes that could potentially bind to an antibody regardless of its type and only a few account for antibody class specific epitope prediction. Recent studies have provided evidence of more than one class of antibodies being associated with a particular disease. Therefore, it is desirable to predict and prioritize ‘peptidome’ representing B-cell epitopes that can potentially bind to multiple classes of antibodies, as an open problem in immunoinformatics. To address this, AbCPE, a novel algorithm based on multi-label classification approach has been developed for prediction of antibody class(es) to which an epitope can potentially bind. The epitopes binding to one or more antibody classes (IgG, IgE, IgA and IgM) have been used as a knowledgebase to derive features for prediction. Multi-label algorithms, Binary Relevance and Label Powerset were applied along with Random Forest and AdaBoost. Classifier performance was assessed using evaluation measures like Hamming Loss, Precision, Recall and F1 score. The Binary Relevance model based on dipeptide composition, Random Forest and AdaBoost achieved the best results with Hamming Loss of 0.1121 and 0.1074 on training and test sets respectively. The results obtained by AbCPE are promising. To the best of our knowledge, this is the first multi-label method developed for prediction of antibody class(es) for sequential B-cell epitopes and is expected to bring a paradigm shift in the field of immunoinformatics and immunotherapeutic developments in synthetic biology. The AbCPE web server is available at http://bioinfo.unipune.ac.in/AbCPE/Home.html.