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• W4311134609 abstract "•IGH genes with high allelic diversity are used in anti-SARS-CoV-2 antibodies•IGH allele usage can influence the activity of neutralizing antibodies•Cryo-EM analysis confirms the role of germline-encoded residues in antigen binding•IGH genotyping can uncover differences in Ab responses due to allelic variation The human immunoglobulin heavy-chain (IGH) locus is exceptionally polymorphic, with high levels of allelic and structural variation. Thus, germline IGH genotypes are personal, which may influence responses to infection and vaccination. For an improved understanding of inter-individual differences in antibody responses, we isolated SARS-CoV-2 spike-specific monoclonal antibodies from convalescent health care workers, focusing on the IGHV1-69 gene, which has the highest level of allelic variation of all IGHV genes. The IGHV1-69∗20-using CAB-I47 antibody and two similar antibodies isolated from an independent donor were critically dependent on allele usage. Neutralization was retained when reverting the V region to the germline IGHV1-69∗20 allele but lost when reverting to other IGHV1-69 alleles. Structural data confirmed that two germline-encoded polymorphisms, R50 and F55, in the IGHV1-69 gene were required for high-affinity receptor-binding domain interaction. These results demonstrate that polymorphisms in IGH genes can influence the function of SARS-CoV-2 neutralizing antibodies. The human immunoglobulin heavy-chain (IGH) locus is exceptionally polymorphic, with high levels of allelic and structural variation. Thus, germline IGH genotypes are personal, which may influence responses to infection and vaccination. For an improved understanding of inter-individual differences in antibody responses, we isolated SARS-CoV-2 spike-specific monoclonal antibodies from convalescent health care workers, focusing on the IGHV1-69 gene, which has the highest level of allelic variation of all IGHV genes. The IGHV1-69∗20-using CAB-I47 antibody and two similar antibodies isolated from an independent donor were critically dependent on allele usage. Neutralization was retained when reverting the V region to the germline IGHV1-69∗20 allele but lost when reverting to other IGHV1-69 alleles. Structural data confirmed that two germline-encoded polymorphisms, R50 and F55, in the IGHV1-69 gene were required for high-affinity receptor-binding domain interaction. These results demonstrate that polymorphisms in IGH genes can influence the function of SARS-CoV-2 neutralizing antibodies. There is an increasing interest in understanding how common variation in immune-related genes influences our ability to control infections, not the least in the context of SARS-CoV-2. As for many other viral infections, inborn deficiencies in type I interferon responses are major risk factors for developing severe coronavirus disease 2019 (COVID-19)2Asano T. Boisson B. Onodi F. Matuozzo D. Moncada-Velez M. Maglorius Renkilaraj M.R.L. Zhang P. Meertens L. Bolze A. Materna M. et al.X-linked recessive TLR7 deficiency in ∼1% of men under 60 years old with life-threatening COVID-19.Sci. Immunol. 2021; 6: eabl4348https://doi.org/10.1126/sciimmunol.abl4348Crossref PubMed Scopus (160) Google Scholar,3Zhang Q. Bastard P. Liu Z. Le Pen J. Moncada-Velez M. Chen J. Ogishi M. Sabli I.K.D. Hodeib S. Korol C. et al.Inborn errors of type I IFN immunity in patients with life-threatening COVID-19.Science. 2020; 370: eabd4570https://doi.org/10.1126/science.abd4570Crossref PubMed Scopus (1239) Google Scholar, while an archaic-derived isoform of oligoadenylate synthetase (OAS), an interferon-induced effector molecule, was shown to confer a protective effect against disease development.4Zeberg H. Pääbo S. A genomic region associated with protection against severe COVID-19 is inherited from Neandertals.Proc. Natl. Acad. Sci. USA. 2021; 118 (e2026309118)https://doi.org/10.1073/pnas.2026309118Crossref PubMed Scopus (102) Google Scholar These and many other findings add to a large body of work describing how variations in the innate immune system influence our response to viruses.5Casanova J.L. Human genetic basis of interindividual variability in the course of infection.Proc. Natl. Acad. Sci. USA. 2015; 112: E7118-E7127https://doi.org/10.1073/pnas.1521644112Crossref PubMed Scopus (123) Google Scholar However, less is known about how variations in adaptive immune receptor genes influence our response to infections.6Peng K. Safonova Y. Shugay M. Popejoy A.B. Rodriguez O.L. Breden F. Brodin P. Burkhardt A.M. Bustamante C. Cao-Lormeau V.M. et al.Diversity in immunogenomics: the value and the challenge.Nat. Methods. 2021; 18: 588-591https://doi.org/10.1038/s41592-021-01169-5Crossref PubMed Scopus (21) Google Scholar The genes that encode the antigen-binding portions of our B cell receptors (BCRs) and T cell receptors (TCRs), the variable (V), diversity (D), and junctional (J) genes, are among the most polymorphic in the human genome, although how this variation imparts functional consequences is largely unknown. The most variable of the three loci that encode the IG genes is the immunoglobulin heavy-chain (IGH) locus, which is present at the telomeric end of chromosome 14. The heavy-chain (HC) locus is characterized by extensive structural variation involving frequent segmental deletions and duplications with particular regions, and the genes contained within, being more affected than others.7Watson C.T. Breden F. The immunoglobulin heavy chain locus: genetic variation, missing data, and implications for human disease.Genes Immun. 2012; 13: 363-373https://doi.org/10.1038/gene.2012.12Crossref PubMed Scopus (114) Google Scholar So far, the number of genomic assemblies, which span a complete human IGH locus, is limited.8Matsuda F. Ishii K. Bourvagnet P. Kuma Ki Hayashida H. Miyata T. Honjo T. The complete nucleotide sequence of the human immunoglobulin heavy chain variable region locus.J. Exp. Med. 1998; 188: 2151-2162https://doi.org/10.1084/jem.188.11.2151Crossref PubMed Scopus (310) Google Scholar,9Rodriguez O.L. Gibson W.S. Parks T. Emery M. Powell J. Strahl M. Deikus G. Auckland K. Eichler E.E. Marasco W.A. et al.A novel framework for characterizing genomic haplotype diversity in the human immunoglobulin heavy chain locus.Front. Immunol. 2020; 11: 2136https://doi.org/10.3389/fimmu.2020.02136Crossref PubMed Scopus (31) Google Scholar,10Watson C.T. Steinberg K.M. Huddleston J. Warren R.L. Malig M. Schein J. Willsey A.J. Joy J.B. Scott J.K. Graves T.A. et al.Complete haplotype sequence of the human immunoglobulin heavy-chain variable, diversity, and joining genes and characterization of allelic and copy-number variation.Am. J. Hum. Genet. 2013; 92: 530-546https://doi.org/10.1016/j.ajhg.2013.03.004Abstract Full Text Full Text PDF PubMed Scopus (155) Google Scholar However, recent studies using BCR repertoire sequencing (Rep-seq) and germline gene inference tools have revealed high inter-individual allelic and structural diversity within the region.9Rodriguez O.L. Gibson W.S. Parks T. Emery M. Powell J. Strahl M. Deikus G. Auckland K. Eichler E.E. Marasco W.A. et al.A novel framework for characterizing genomic haplotype diversity in the human immunoglobulin heavy chain locus.Front. Immunol. 2020; 11: 2136https://doi.org/10.3389/fimmu.2020.02136Crossref PubMed Scopus (31) Google Scholar,11Corcoran M.M. Phad G.E. Vázquez Bernat N. Stahl-Hennig C. Sumida N. Persson M.A. Martin M. Karlsson Hedestam G.B. Production of individualized V gene databases reveals high levels of immunoglobulin genetic diversity.Nat. Commun. 2016; 7: 13642https://doi.org/10.1038/ncomms13642Crossref PubMed Scopus (116) Google Scholar,12Gidoni M. Snir O. Peres A. Polak P. Lindeman I. Mikocziova I. Sarna V.K. Lundin K.E.A. Clouser C. Vigneault F. et al.Mosaic deletion patterns of the human antibody heavy chain gene locus shown by Bayesian haplotyping.Nat. Commun. 2019; 10: 628https://doi.org/10.1038/s41467-019-08489-3Crossref PubMed Scopus (46) Google Scholar Importantly, IGHV allele usage determines the initial states of the HC complementarity determining regions 1 and 2 (HCDR1 and HCDR2) and contributes to the HCDR3, all of which shape epitope binding. The degree to which the presence or absence of specific IGH alleles influences the development of antibody responses to different pathogens remains largely unknown. To date, only a few examples of allele-specific responses have been described. These include a requirement for IGHV1-2∗02 or IGHV1-2∗04 for the generation of VRC01 class neutralizing antibodies directed against the CD4 binding site of HIV-113Lee J.H. Toy L. Kos J.T. Safonova Y. Schief W.R. Havenar-Daughton C. Watson C.T. Crotty S. Vaccine genetics of IGHV1-2 VRC01-class broadly neutralizing antibody precursor naïve human B cells.npj Vaccines. 2021; 6: 113https://doi.org/10.1038/s41541-021-00376-7Crossref PubMed Scopus (19) Google Scholar and the propensity of specific IGHV1-69 alleles to be utilized in influenza hemagglutinin (HA) stem-directed neutralizing antibodies.14Avnir Y. Tallarico A.S. Zhu Q. Bennett A.S. Connelly G. Sheehan J. Sui J. Fahmy A. Huang C.Y. Cadwell G. et al.Molecular signatures of hemagglutinin stem-directed heterosubtypic human neutralizing antibodies against influenza A viruses.PLoS Pathog. 2014; 10: e1004103https://doi.org/10.1371/journal.ppat.1004103Crossref PubMed Scopus (93) Google Scholar,15Avnir Y. Watson C.T. Glanville J. Peterson E.C. Tallarico A.S. Bennett A.S. Qin K. Fu Y. Huang C.Y. Beigel J.H. et al.IGHV1-69 polymorphism modulates anti-influenza antibody repertoires, correlates with IGHV utilization shifts and varies by ethnicity.Sci. Rep. 2016; 6: 20842https://doi.org/10.1038/srep20842Crossref PubMed Scopus (107) Google Scholar Previous SARS-CoV-2 antibody studies demonstrated that certain IGHV genes are frequently used among S-specific neutralizing antibodies, including IGHV1-69, IGHV3-30, IGHV3-30-3, IGHV3-53, IGHV3-66, and IGHV5-51.16Claireaux M.e.a. A public antibody class recognizes a novel S2 epitope exposed on open conformations of SARS-CoV-2 spike.Preprint at bioRxiv. 2022; https://doi.org/10.1101/2021.12.01.470767v1Crossref Google Scholar,17Robbiani D.F. Gaebler C. Muecksch F. Lorenzi J.C.C. Wang Z. Cho A. Agudelo M. Barnes C.O. Gazumyan A. Finkin S. et al.Convergent antibody responses to SARS-CoV-2 in convalescent individuals.Nature. 2020; 584: 437-442https://doi.org/10.1038/s41586-020-2456-9Crossref PubMed Scopus (1122) Google Scholar,18Sakharkar M. Rappazzo C.G. Wieland-Alter W.F. Hsieh C.L. Wrapp D. Esterman E.S. Kaku C.I. Wec A.Z. Geoghegan J.C. McLellan J.S. et al.Prolonged evolution of the human B cell response to SARS-CoV-2 infection.Sci. Immunol. 2021; 6: eabg6916https://doi.org/10.1126/sciimmunol.abg6916Crossref PubMed Google Scholar,19Yuan M. Liu H. Wu N.C. Wilson I.A. Recognition of the SARS-CoV-2 receptor binding domain by neutralizing antibodies.Biochem. Biophys. Res. Commun. 2021; 538: 192-203https://doi.org/10.1016/j.bbrc.2020.10.012Crossref PubMed Scopus (111) Google Scholar Of these, IGHV1-69 is of particular interest as it has the highest number of known allelic variants and is associated with structural variations resulting in frequent gene duplications.9Rodriguez O.L. Gibson W.S. Parks T. Emery M. Powell J. Strahl M. Deikus G. Auckland K. Eichler E.E. Marasco W.A. et al.A novel framework for characterizing genomic haplotype diversity in the human immunoglobulin heavy chain locus.Front. Immunol. 2020; 11: 2136https://doi.org/10.3389/fimmu.2020.02136Crossref PubMed Scopus (31) Google Scholar,10Watson C.T. Steinberg K.M. Huddleston J. Warren R.L. Malig M. Schein J. Willsey A.J. Joy J.B. Scott J.K. Graves T.A. et al.Complete haplotype sequence of the human immunoglobulin heavy-chain variable, diversity, and joining genes and characterization of allelic and copy-number variation.Am. J. Hum. Genet. 2013; 92: 530-546https://doi.org/10.1016/j.ajhg.2013.03.004Abstract Full Text Full Text PDF PubMed Scopus (155) Google Scholar,15Avnir Y. Watson C.T. Glanville J. Peterson E.C. Tallarico A.S. Bennett A.S. Qin K. Fu Y. Huang C.Y. Beigel J.H. et al.IGHV1-69 polymorphism modulates anti-influenza antibody repertoires, correlates with IGHV utilization shifts and varies by ethnicity.Sci. Rep. 2016; 6: 20842https://doi.org/10.1038/srep20842Crossref PubMed Scopus (107) Google Scholar,20Luo S. Yu J.A. Song Y.S. Estimating copy number and allelic variation at the immunoglobulin heavy chain locus using short reads.PLoS Comput. Biol. 2016; 12: e1005117https://doi.org/10.1371/journal.pcbi.1005117Crossref PubMed Scopus (16) Google Scholar Critically, IGHV1-69 is among the most highly utilized IGHV genes in the naive B cell repertoire.12Gidoni M. Snir O. Peres A. Polak P. Lindeman I. Mikocziova I. Sarna V.K. Lundin K.E.A. Clouser C. Vigneault F. et al.Mosaic deletion patterns of the human antibody heavy chain gene locus shown by Bayesian haplotyping.Nat. Commun. 2019; 10: 628https://doi.org/10.1038/s41467-019-08489-3Crossref PubMed Scopus (46) Google Scholar Therefore, there is a high probability that B cells using this gene will encounter antigens after infection. However, because of the high allelic variation, antibody responses involving IGHV1-69 may differ considerably between individuals. Importantly, studies describing S-specific monoclonal antibodies (mAbs) isolated from convalescent individuals early after exposure showed that few somatic hypermutations (SHMs) are sufficient to confer neutralizing activity against SARS-CoV-2.21Brouwer P.J.M. Caniels T.G. van der Straten K. Snitselaar J.L. Aldon Y. Bangaru S. Torres J.L. Okba N.M.A. Claireaux M. Kerster G. et al.Potent neutralizing antibodies from COVID-19 patients define multiple targets of vulnerability.Science. 2020; 369: 643-650https://doi.org/10.1126/science.abc5902Crossref PubMed Scopus (677) Google Scholar,22Kreer C. Zehner M. Weber T. Ercanoglu M.S. Gieselmann L. Rohde C. Halwe S. Korenkov M. Schommers P. Vanshylla K. et al.Longitudinal isolation of potent near-germline SARS-CoV-2-neutralizing antibodies from COVID-19 patients.Cell. 2020; 182: 843-854.e12https://doi.org/10.1016/j.cell.2020.06.044Abstract Full Text Full Text PDF PubMed Scopus (180) Google Scholar,23Seydoux E. Homad L.J. MacCamy A.J. Parks K.R. Hurlburt N.K. Jennewein M.F. Akins N.R. Stuart A.B. Wan Y.H. Feng J. et al.Analysis of a SARS-CoV-2-infected individual reveals development of potent neutralizing antibodies with limited somatic mutation.Immunity. 2020; 53: 98-105.e5https://doi.org/10.1016/j.immuni.2020.06.001Abstract Full Text Full Text PDF PubMed Scopus (228) Google Scholar Many of these antibodies target the receptor-binding domain (RBD) of the spike trimer, and research has also shown that RBD-binding B cells can be isolated from SARS-CoV-2 seronegative individuals.24Feldman J. Bals J. Altomare C.G. St Denis K. Lam E.C. Hauser B.M. Ronsard L. Sangesland M. Moreno T.B. Okonkwo V. et al.Naive human B cells engage the receptor binding domain of SARS-CoV-2, variants of concern, and related sarbecoviruses.Sci. Immunol. 2021; 6: eabl5842https://doi.org/10.1126/sciimmunol.abl5842Crossref PubMed Scopus (16) Google Scholar If unmutated or modestly mutated antibodies that use IGHV genes with high allelic variation display neutralizing activity, it follows that this variation could impact the quality of both the initial and subsequent humoral response to the virus. To address this question, we carried out personalized IG genotyping of previously infected healthcare workers to identify allelic and structural variations, particularly those affecting IGHV1-69. We first selected a case with high IGHV allelic content and a high frequency of circulating S-specific memory B cells from whom we isolated and characterized a set of SARS-CoV-2 S-specific mAbs (n = 29). Individualized genotyping enabled us to assign all cloned antibodies to the specific IGHV, IGHD, and IGHJ alleles present in this person. Five of the mAbs that we found to be potently neutralizing used the IGHV1-69 gene, of which three, CAB-I12, CAB-I47, and CAB-J39, were assigned to a recently described IGHV1-69∗20 allele.12Gidoni M. Snir O. Peres A. Polak P. Lindeman I. Mikocziova I. Sarna V.K. Lundin K.E.A. Clouser C. Vigneault F. et al.Mosaic deletion patterns of the human antibody heavy chain gene locus shown by Bayesian haplotyping.Nat. Commun. 2019; 10: 628https://doi.org/10.1038/s41467-019-08489-3Crossref PubMed Scopus (46) Google Scholar We investigated the impact of IGHV allele usage by reverting one of the most potent IGHV1-69∗20 using antibodies, CAB-I47 to the IGHV1-69∗20 germline sequence and to an additional set of five IGHV1-69 alleles that are common in the global human population. Assessment of the neutralizing activities of these antibody versions revealed that allele usage was critical for functional activity. Two additional IGHV1-69∗20-using neutralizing antibodies, CAB-M77 and CAB-N86, which were isolated from an independent donor, showed very similar allele dependence. A high-resolution cryo-EM structure of the CAB-I47 Fab bound to the spike, revealed the molecular details of its interaction with the RBD. Overall, our results demonstrate that IGHV allele usage can profoundly influence the neutralizing antibody response after SARS-CoV-2 infection. Further studies aimed at defining how germline gene variation influences anti-viral B cell responses at individual level and population levels will improve our understanding of adaptive immunity. A cohort of healthcare workers (n = 14), who tested positive for SARS-CoV-2 by RT-PCR in May 2020 was established to study the evolution of S-specific antibodies and memory B cell responses. Peripheral blood mononuclear cells (PBMCs) and serum samples were collected 7 months after primary infection and used for serological analyses, IG genotyping, and mAb isolation (Figure 1A). All study participants (SP01–SP14) were seropositive for S-IgG (Figure 1B), and donors displayed detectable neutralizing activity at this time point (Figure 1C). To determine the IGHV genotypes of the participants, we produced libraries from the expressed IgM repertoire from the same time point, bulk sequenced these using the MiSeq platform, and analyzed them using our germline allele inference tool, IgDiscover.11Corcoran M.M. Phad G.E. Vázquez Bernat N. Stahl-Hennig C. Sumida N. Persson M.A. Martin M. Karlsson Hedestam G.B. Production of individualized V gene databases reveals high levels of immunoglobulin genetic diversity.Nat. Commun. 2016; 7: 13642https://doi.org/10.1038/ncomms13642Crossref PubMed Scopus (116) Google Scholar The results revealed high IGHV diversity between individuals, both in terms of the number of alleles, ranging from 44 to 61 per person, and the content. The complete IGHV, IGHD, and IGHJ genotypes are shown in Table S1, demonstrating that the number of IGHV1-69 alleles varied greatly between the study participants. One individual was homozygous for IGHV1-69∗01, while most study participants had two or three different alleles and two individuals had as many as four different IGHV1-69 alleles. Amino acid sequence alignment of all functional IGHV1-69 alleles shows that there are six variant positions between the alleles (Figure 1D). Altogether, six different alleles were found in the 14 study participants, IGHV1-69∗01, IGHV1-69∗02, IGHV1-69∗04, IGHV1-69∗06, IGHV1-69∗09, and IGHV1-69∗20 (Figure 1E). To determine which alleles were present on each chromosome in the study participants, we performed inferred haplotype analysis.25Kirik U. Greiff L. Levander F. Ohlin M. Parallel antibody germline gene and haplotype analyses support the validity of immunoglobulin germline gene inference and discovery.Mol. Immunol. 2017; 87: 12-22https://doi.org/10.1016/j.molimm.2017.03.012Crossref PubMed Scopus (26) Google Scholar,26Vázquez Bernat N. Corcoran M. Nowak I. Kaduk M. Castro Dopico X. Narang S. Maisonasse P. Dereuddre-Bosquet N. Murrell B. Karlsson Hedestam G.B. Rhesus and cynomolgus macaque immunoglobulin heavy-chain genotyping yields comprehensive databases of germline VDJ alleles.Immunity. 2021; 54: 355-366.e4https://doi.org/10.1016/j.immuni.2020.12.018Abstract Full Text Full Text PDF PubMed Scopus (24) Google Scholar This method takes advantage of the fact that VDJ recombination occurs locally on a given chromosome and heterozygous IGHJ or IGHD alleles as anchors to link IGHV alleles to either chromosome. Examining IGHD and IGHJ allele content, we found that SP01, SP02, SP06, SP08, and SP11 were heterozygous for IGHJ (IGHJ6∗02/IGHJ6∗03), SP14 was heterozygous for IGHD2-21 (IGHD2-21∗01/IGHD2-21∗02), and SP04 and SP05 were heterozygous for IGHD3-10 (IGHD3-10∗01/IGHD3-10∗01_S2851) (Figures 1F and S1A). Use of these anchors to haplotype IGHV alleles demonstrated that the presence of gene duplications resulting in structural variation between the chromosomes (Figures 1F and S1A). Overall, the haplotype analysis of eight individuals in this cohort confirmed high allelic and structural variation in the IGHV1-69 region of the IGH locus. To expand upon this in a larger cohort, we analyzed available IgM repertoire data from 32 unrelated cases not previously infected with SARS-CoV-2.12Gidoni M. Snir O. Peres A. Polak P. Lindeman I. Mikocziova I. Sarna V.K. Lundin K.E.A. Clouser C. Vigneault F. et al.Mosaic deletion patterns of the human antibody heavy chain gene locus shown by Bayesian haplotyping.Nat. Commun. 2019; 10: 628https://doi.org/10.1038/s41467-019-08489-3Crossref PubMed Scopus (46) Google Scholar In this dataset, IGHV1-69 was the third most frequently used gene in the IgM repertoire (Figure S1B). Of the 32 individuals examined, nine could be haplotyped using heterozygous IGHJ6 as an anchor. This revealed that three of these cases had more than two alleles and one study participant had four IGHV1-69 alleles (Figure S1B). Together, these data demonstrated high allelic and structural variation in the IGHV1-69 region within the populations studied. We selected one of the participants from the SARS-CoV-2 convalescent cohort, SP14, who had three IGHV1-69 alleles (IGHV1-69∗01, IGHV1-69∗02, and IGHV1-69∗20) (Figure 2A). The participant’s kappa and lambda light-chain (LC) V germline alleles, IGKV and IGLV, were also determined (Figure 2A). Single SARS-CoV-2 S-specific IgG+ memory B cells were sorted into 96-well plates for antibody sequence analysis and mAb isolation. Of the CD20+CD27+IgG+ memory B cells, 2.7% were S specific (Figures 2B and S2A). Amplification of antibody V(D)J regions from the 247 sorted cells was performed by single-cell RT-PCR using a previously published comprehensive primer set for human HC and LC isolation.27Vázquez Bernat N. Corcoran M. Hardt U. Kaduk M. Phad G.E. Martin M. Karlsson Hedestam G.B. High-quality library preparation for NGS-based immunoglobulin germline gene inference and repertoire expression analysis.Front. Immunol. 2019; 10: 660https://doi.org/10.3389/fimmu.2019.00660Crossref PubMed Scopus (29) Google Scholar Sanger sequencing identified 177 full-length HC sequences and 146 paired HC and LC sequences, a subset of which were cloned into expression vectors28Tiller T. Meffre E. Yurasov S. Tsuiji M. Nussenzweig M.C. Wardemann H. Efficient generation of monoclonal antibodies from single human B cells by single cell RT-PCR and expression vector cloning.J. Immunol. Methods. 2008; 329: 112-124https://doi.org/10.1016/j.jim.2007.09.017Crossref PubMed Scopus (698) Google Scholar to produce mAbs. Examination of the IGHV allele usage of the 177 spike-specific HCs and comparisons with the study participant’s total IgG repertoire, obtained by bulk PBMC RNA sequencing, revealed that several IGHV genes were over-represented in the S-specific B cells, including IGHV1-69, IGHV3-30-3, IGHV3-53, and IGHV5-51, consistent with previous reports19Yuan M. Liu H. Wu N.C. Wilson I.A. Recognition of the SARS-CoV-2 receptor binding domain by neutralizing antibodies.Biochem. Biophys. Res. Commun. 2021; 538: 192-203https://doi.org/10.1016/j.bbrc.2020.10.012Crossref PubMed Scopus (111) Google Scholar,29Barnes C.O. Jette C.A. Abernathy M.E. Dam K.A. Esswein S.R. Gristick H.B. Malyutin A.G. Sharaf N.G. Huey-Tubman K.E. Lee Y.E. et al.SARS-CoV-2 neutralizing antibody structures inform therapeutic strategies.Nature. 2020; 588: 682-687https://doi.org/10.1038/s41586-020-2852-1Crossref PubMed Scopus (770) Google Scholar,30Tan T.J.C. Yuan M. Kuzelka K. Padron G.C. Beal J.R. Chen X. Wang Y. Rivera-Cardona J. Zhu X. Stadtmueller B.M. et al.Sequence signatures of two public antibody clonotypes that bind SARS-CoV-2 receptor binding domain.Nat. Commun. 2021; 12: 3815https://doi.org/10.1038/s41467-021-24123-7Crossref PubMed Scopus (21) Google Scholar,31Vanshylla K. Fan C. Wunsch M. Poopalasingam N. Meijers M. Kreer C. Kleipass F. Ruchnewitz D. Ercanoglu M.S. Gruell H. et al.Discovery of ultrapotent broadly neutralizing antibodies from SARS-CoV-2 elite neutralizers.Cell Host Microbe. 2022; 30: 69-82.e10https://doi.org/10.1016/j.chom.2021.12.010Abstract Full Text Full Text PDF PubMed Scopus (19) Google Scholar,32Yuan M. Liu H. Wu N.C. Lee C.D. Zhu X. Zhao F. Huang D. Yu W. Hua Y. Tien H. et al.Structural basis of a shared antibody response to SARS-CoV-2.Science. 2020; 369: 1119-1123https://doi.org/10.1126/science.abd2321Crossref PubMed Scopus (320) Google Scholar,33Zhou X. Ma F. Xie J. Yuan M. Li Y. Shaabani N. Zhao F. Huang D. Wu N.C. Lee C.D. et al.Diverse immunoglobulin gene usage and convergent epitope targeting in neutralizing antibody responses to SARS-CoV-2.Cell Rep. 2021; 35: 109109https://doi.org/10.1016/j.celrep.2021.109109Abstract Full Text Full Text PDF PubMed Scopus (13) Google Scholar,34Zost S.J. Gilchuk P. Chen R.E. Case J.B. Reidy J.X. Trivette A. Nargi R.S. Sutton R.E. Suryadevara N. Chen E.C. et al.Rapid isolation and profiling of a diverse panel of human monoclonal antibodies targeting the SARS-CoV-2 spike protein.Nat. Med. 2020; 26: 1422-1427https://doi.org/10.1038/s41591-020-0998-xCrossref PubMed Scopus (268) Google Scholar (Figure 2C). The S-specific response was highly polyclonal, with 171 unique clones of the 177 total HC sequences analyzed (Figure 2D) using the same IGHV allele, IGHJ allele, length of HCDR3, and 80% aa HCDR3 identity as a definition for clonal relatedness.35Phad G.E. Pushparaj P. Tran K. Dubrovskaya V. Àdori M. Martinez-Murillo P. Vázquez Bernat N. Singh S. Dionne G. O'Dell S. et al.Extensive dissemination and intraclonal maturation of HIV Env vaccine-induced B cell responses.J. Exp. Med. 2020; 217: e20191155https://doi.org/10.1084/jem.20191155Crossref PubMed Google Scholar From the 146 paired sequences, 29 spike-specific mAbs were expressed, of which 52% (n = 15) were neutralizing (Figure 2E). All neutralizing mAbs bound the RBD (Figure S2B). The neutralization curves and IC50 values against the ancestral SARS-CoV-2 strain are shown in Figure S2C. A summary of the genetic and functional properties of all neutralizing mAbs is shown in Figure 2F and of the non-neutralizing mAbs in Table S2. Knowledge of the specific V, D, and J alleles present in the study participant allowed for precise genetic characterization of the isolated neutralizing antibodies. Several mAbs used IGHV3-53 and had short HCDR3s, suggesting that they were of the previously described class 1 group of SARS-CoV-2-specific antibodies.32Yuan M. Liu H. Wu N.C. Lee C.D. Zhu X. Zhao F. Huang D. Yu W. Hua Y. Tien H. et al.Structural basis of a shared antibody response to SARS-CoV-2.Science. 2020; 369: 1119-1123https://doi.org/10.1126/science.abd2321Crossref PubMed Scopus (320) Google Scholar,36Barnes C.O. West Jr., A.P. Huey-Tubman K.E. Hoffmann M.A.G. Sharaf N.G. Hoffman P.R. Koranda N. Gristick H.B. Gaebler C. Muecksch F. et al.Structures of human antibodies bound to SARS-CoV-2 spike reveal common epitopes and recurrent features of antibodies.Cell. 2020; 182: 828-842.e16https://doi.org/10.1016/j.cell.2020.06.025Abstract Full Text Full Text PDF PubMed Scopus (448) Google Scholar The IGHV3-53∗01-using mAbs isolated here, including three clonally related mAbs, CAB-A17, CAB-A18, and CAB-A49, are separately described.37Sheward D.J.P. P. Das H. Kim C. Kim S. Hanke L. Dyrdak R. McInerney G. Albert A. Murrell B. et al.Structural basis of Omicron neutralization by affinity-matured public antibodies.Preprint at bioRxiv. 2022; https://doi.org/10.1101/2022.01.03.474825v1Crossref Google Scholar One ultrapotent neutralizing mAb, CAB-F52, used IGHV3-30-3∗01, a gene that is present in only a subset of individuals (Table S1). Several of the most potently neutralizing mAbs used IGHV1-69, either IGHV1-69∗02 or the recently identified IGHV1-69∗20 allele.12Gidoni M. Snir O. Peres A. Polak P. Lindeman I. Mikocziova I. Sarna V.K. Lundin K.E.A. Clouser C. Vigneault F. et al.Mosaic deletion patterns of the human antibody heavy chain gene locus shown by Bayesian haplotyping.Nat. Commun. 2019; 10: 628https://doi.org/10.1038/s41467-019-08489-3Crossref PubMed Scopus (46) Google Scholar Two of the IGHV1-69∗20-using mAbs, CAB-I12 and CAB-I47, were clonally related (Figure 2F) and narrow in their neutralization profile against different SARS-CoV-2 variants (Figure S2D). When inspecting the sequence alignment of the IGHV1-69 alleles, we found that IGHV1-69∗20 was closest to IGHV1-69∗04 with a single-nucleotide difference resulting in a leucine (L) in ∗04 to phenylalanine (F) substitution at position 55 in the HCDR2 of ∗20. Several other IGHV1-69 alleles, ∗01, ∗05, ∗06, ∗12, ∗13, ∗15, and ∗17, also have an F in this position but differ from IGHV1-69∗20 at other variant positions (Figure 1D). The average SHM for the neutralizing mAbs was 5.5% in the nucleotide sequence and 10.1% in the aa sequence, reflecting the time point at which they were isolated, 7 months after primary infection. None of the mAbs used previously unreported LC alleles. These results demonstrate that potent" @default.
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• W4311134609 title "Immunoglobulin germline gene polymorphisms influence the function of SARS-CoV-2 neutralizing antibodies" @default.
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• W4311134609 doi "https://doi.org/10.1016/j.immuni.2022.12.005" @default.
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