9 research outputs found

    Persistent antibody clonotypes dominate the serum response to influenza following repeated vaccination over multiple years

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    We used Ig-Seq, a liquid chromatography tandem mass spectrometry (LC-MS/MS)–based serum antibody proteomics methodology, to determine the clonal composition and dynamics of the H1N1 California/7/2009 (CA09) hemagglutinin (HA)-reactive antibody repertoire over 5 years in a well-characterized donor from whom a large number of homosubtypic and heterosubtypic neutralizing monoclonal antibodies had been previously isolated by B cell analysis. The donor was infected with the CA09 strain in 2009 and immunized annually for the next five years with seasonal influenza vaccine which contained the CA09 strain. We find that the serological repertoire in this donor was highly static, with a modest number (24) of persistent antibody clonotypes, detected in serum for at least 4 out of 5 years, accounting on average for 72.6 ± 10.0% of the repertoire to the CA09 HA. These persistent antibodies: (i) displayed a higher degree of somatic hypermutation relative to antibodies that could be detected in the serum transiently (i.e. lasted less than 1 year in serum); (ii) comprised a significant fraction that also bound to HA from a phylogenetically distant H5N1 A/Vietnam/1203/2004 (VT04) strain, a hallmark of stem-binding antibodies due to the lack of homology between CA09 and VT04 in the head region of HA and (iii) perhaps most strikingly, but consistent with the wealth of heterosubtypic neutralizing antibodies that had previously been identified from this donor, some of the most abundant persistent antibody clonotypes, including the dominant clone that accounted on average for 18.6 ± 12.3% of the serum titer across 5 years, neutralized both the CA09 and VT04 influenza strains. Our analysis highlights the magnitude of ‘serological imprinting’ in the donor’s serum response to CA09, indicates that seasonal vaccination can further reinforce a stable serological memory and finally suggests that once elicited, antibodies cross-reactive between CA09 and VT04 with heterosubtypic neutralization activity, thus likely to bind to HA-stem, can persist for many years, which is a fundamental goal of universal influenza vaccines

    ACE2-binding exposes the SARS-CoV-2 fusion peptide to broadly neutralizing coronavirus antibodies

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    The coronavirus spike glycoprotein attaches to host receptors and mediates viral fusion. Using a broad screening approach, we isolated seven monoclonal antibodies (mAbs) that bind to all human-infecting coronavirus spike proteins from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immune donors. These mAbs recognize the fusion peptide and acquire affinity and breadth through somatic mutations. Despite targeting a conserved motif, only some mAbs show broad neutralizing activity in vitro against alpha- and betacoronaviruses, including animal coronaviruses WIV-1 and PDF-2180. Two selected mAbs also neutralize Omicron BA.1 and BA.2 authentic viruses and reduce viral burden and pathology in vivo. Structural and functional analyses showed that the fusion peptide–specific mAbs bound with different modalities to a cryptic epitope hidden in prefusion stabilized spike, which became exposed upon binding of angiotensin-converting enzyme 2 (ACE2) or ACE2-mimicking mAbs

    Bioinformatic analyses of integral membrane transport proteins encoded within the genome of the planctomycetes species, Rhodopirellula baltica

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    Rhodopirellula baltica (R. baltica) is a Planctomycete, known to have intracellular membranes. Because of its unusual cell structure and ecological significance, we have conducted comprehensive analyses of its transmembrane transport proteins. The complete proteome of R. baltica was screened against the Transporter Classification Database (TCDB) to identify recognizable integral membrane transport proteins. 342 proteins were identified with a high degree of confidence, and these fell into several different classes. R. baltica encodes in its genome channels (12%), secondary carriers (33%), and primary active transport proteins (41%) in addition to classes represented in smaller numbers. Relative to most non-marine bacteria, R. baltica possesses a larger number of sodium-dependent symporters but fewer proton-dependent symporters, and it has dimethylsulfoxide (DMSO) and trimethyl-amine-oxide (TMAO) reductases, consistent with its Na-rich marine environment. R. baltica also possesses a Na-translocating NADH:quinone dehydrogenase (Na-NDH), a Na efflux decarboxylase, two Na-exporting ABC pumps, two Na-translocating F-type ATPases, two Na:H antiporters and two K:H antiporters. Flagellar motility probably depends on the sodium electrochemical gradient. Surprisingly, R. baltica also has a complete set of H-translocating electron transport complexes similar to those present in α-proteobacteria and eukaryotic mitochondria. The transport proteins identified proved to be typical of the bacterial domain with little or no indication of the presence of eukaryotic-type transporters. However, novel functionally uncharacterized multispanning membrane proteins were identified, some of which are found only in Rhodopirellula species, but others of which are widely distributed in bacteria. The analyses lead to predictions regarding the physiology, ecology and evolution of R. baltica

    Deciphering and predicting CD4+ T cell immunodominance of influenza virus hemagglutinin

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    The importance of CD4+ T helper (Th) cells is well appreciated in view of their essential role in the elicitation of antibody and cytotoxic T cell responses. However, the mechanisms that determine the selection of immunodominant epitopes within complex protein antigens remain elusive. Here, we used ex vivo stimulation of memory T cells and screening of naive and memory T cell libraries, combined with T cell cloning and TCR sequencing, to dissect the human naive and memory CD4+ T cell repertoire against the influenza pandemic H1 hemagglutinin (H1-HA). We found that naive CD4+ T cells have a broad repertoire, being able to recognize naturally processed as well as cryptic peptides spanning the whole H1-HA sequence. In contrast, memory Th cells were primarily directed against just a few immunodominant peptides that were readily detected by mass spectrometry–based MHC-II peptidomics and predicted by structural accessibility analysis. Collectively, these findings reveal the presence of a broad repertoire of naive T cells specific for cryptic H1-HA peptides and demonstrate that antigen processing represents a major constraint determining immunodominance.ISSN:0022-1007ISSN:1540-0069ISSN:1540-953

    ACE2-binding exposes the SARS-CoV-2 fusion peptide to broadly neutralizing coronavirus antibodies

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    The coronavirus spike (S) glycoprotein attaches to host receptors and mediates viral fusion. Using a broad screening approach, we isolated from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immune donors seven monoclonal antibodies (mAbs) that bind to all human-infecting coronavirus S proteins. This class of mAbs recognize the fusion peptide and acquire affinity and breadth through somatic mutations. Despite targeting a conserved motif, only some mAbs show broad neutralizing activity in vitro against alpha- and beta-coronaviruses, including animal coronavirus WIV-1 and PDF-2180. Two selected mAbs also neutralize Omicron BA.1 and BA.2 authentic viruses and reduce viral burden and pathology in vivo. Structural and functional analyses show that the fusion peptide-specific mAbs bind with different modalities to a cryptic epitope, which is hidden in prefusion stabilized S, and becomes exposed upon binding of angiotensin-converting enzyme 2 (ACE2) or ACE2-mimicking mAbs.ISSN:0036-8075ISSN:1095-920

    Identification and structure of a multidonor class of head-directed influenza-neutralizing antibodies reveal the mechanism for its recurrent elicitation

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    Multidonor antibodies are of interest for vaccine design because they can in principle be elicited in the general population by a common set of immunogens. For influenza, multidonor antibodies have been observed against the hemagglutinin (HA) stem, but not the immunodominant HA head. Here, we identify and characterize a multidonor antibody class (LPAF-a class) targeting the HA head. This class exhibits potent viral entry inhibition against H1N1 A/California/04/2009 (CA09) virus. LPAF-a class antibodies derive from the HV2-70 gene and contain a “Tyr-Gly-Asp”-motif, which occludes the HA-sialic acid binding site as revealed by a co-crystal structure with HA. Both germline-reverted and mature LPAF antibodies potently neutralize CA09 virus and have nanomolar affinities for CA09 HA. Moreover, increased frequencies for LPFA-a class antibodies are observed in humans after a single vaccination. Overall, this work highlights the identification of a multidonor class of head-directed influenza-neutralizing antibodies and delineates the mechanism of their recurrent elicitation in humans

    Deciphering and predicting CD4+ T cell immunodominance of influenza virus hemagglutinin

    No full text
    The importance of CD4+ T helper (Th) cells is well appreciated in view of their essential role in the elicitation of antibody and cytotoxic T cell responses. However, the mechanisms that determine the selection of immunodominant epitopes within complex protein antigens remain elusive. Here, we used ex vivo stimulation of memory T cells and screening of naive and memory T cell libraries, combined with T cell cloning and TCR sequencing, to dissect the human naive and memory CD4+ T cell repertoire against the influenza pandemic H1 hemagglutinin (H1-HA). We found that naive CD4+ T cells have a broad repertoire, being able to recognize naturally processed as well as cryptic peptides spanning the whole H1-HA sequence. In contrast, memory Th cells were primarily directed against just a few immunodominant peptides that were readily detected by mass spectrometry–based MHC-II peptidomics and predicted by structural accessibility analysis. Collectively, these findings reveal the presence of a broad repertoire of naive T cells specific for cryptic H1-HA peptides and demonstrate that antigen processing represents a major constraint determining immunodominance.ISSN:0022-1007ISSN:1540-0069ISSN:1540-953
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