bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sequential and interwoven requirements for HEB and Id3 in fetal gamma-delta T cell commitment and functional programming
Authors: Anderson Michele K , Selvaratnam Johanna , Rocha Juliana , Rajan Vinothkumar , Wang Helen , Reddy Emily , Guidos Cynthia , Zuniga-Pflucker Juan Carlos 
Published: August 2023

Abstract
Two major developmental steps are necessary for the generation of innate IL-17 producing γδ T cells. First, T cell precursors in the fetal thymus commit to either the αβ or the γδ T cell lineage. This is followed by a second stage of differentiation into subsets that can produce either IL-17 (γδT17) or IFNγ (γδT1) cells. Both developmental events are dependent on TCR signal strength, but how this is interpreted at the molecular level has been unclear. Id3, which antagonizes the activity of HEB transcription factors, is upregulated in proportion to TCR signal strength, providing an important link between TCR signaling and gene expression. We previously found that γδ T cells in mice lacking HEB were impaired in IL-17 production. To better understand the molecular basis of this defect, we conducted single cell RNA-sequencing on fetal thymic γδ T cells. In WT cells, the ratio of HEB to Id3 was highest in populations undergoing γδ T cell commitment, whereas later populations were dominated by Id3. Comparing scRNA-seq datasets from WT and HEB-deficient γδ T cells revealed a profound shift in TCRγ and TCRδ chain expression. This was accompanied by a decrease in key regulators of early γδ T cell development. Id3 expression was also severely decreased. Accordingly, Id3-deficient mice were impaired in their ability to generate γδT17 cells. However, γδ T cells from Id3-deficient mice showed a decrease in second stage γδT17 regulators, rather than the first stage regulators defective in HEB-deficient mice. Therefore, distinct ratios of HEB and Id3 are required during the γδ T cell commitment, and HEB is instrumental in inducing Id3 to enable functional programming of γδT17 cells in the fetal thymus.
Supported by grants from CIHR (201610PJT), NSERC (RGPIN-2020-05596) and NIH (1P01AI102853-06)
The ER aminopeptidases, ERAP1 and ERAP2, Synergize to Self-Modulate their Respective Activities
Authors: Martin-Esteban Adrian , Rodriguez Jesus Contreras , Peske David J , de Castro Jose Antonio Lopez , Shastri Nilabh , Sadegh-Nasseri Scheherazade 
Published: August 2023

Abstract
Critical steps in Major Histocompatibility Complex Class I (MHC-I) antigen presentation occur in the endoplasmic reticulum (ER). In general, peptides that enter the ER are longer than the optimal length for MHC-I binding. The final trimming of MHC-I epitopes is performed by two related aminopeptidases, ERAP1 and ERAP2 in humans that possess unique and complementary substrate trimming specificities. While ERAP1 efficiently trims peptides longer than 9 residues, ERAP2 preferentially trims peptides shorter than 9 residues. Using a combination of biochemical and proteomic studies followed by biological verification, we demonstrate that the optimal ligands for either enzyme act as inhibitors of the other enzyme. Specifically, the presence of octamers reduced the trimming of long peptides by ERAP1, while peptides longer than nonomers inhibit ERAP2 activity. We propose a mechanism for how ERAP1 and ERAP2 synergize to modulate their respective activities and shape the MHC-I peptidome by generating optimal peptides for presentation.
This work was supported by grants ROI AI121174, R-37AI-060040, ROI AI-130210 from the National Institutes of Health (NIH) and SAF2017-86578-R from the Ministry of Science, Innovation, and Universities of the Government of Spain and by an institutional grant of the Fundación Ramón Areces to the CBMSO.
Recombinant Surface Immunogenic Protein from S. agalactiae and hemocyanin from Fisurella latimarginata promote cross-presentation dependence on the TLR-4 signaling
Authors: Diaz-Dinamarca Diego A , Salazar Michelle , Manubens Augusto , Salazar Fabián , Troncoso Mayarling F , Lavandero Sergio , Diaz Janepsy , Vasquez Abel E , Becker María Inés 
Published: August 2023

Abstract
Improved vaccine adjuvants are needed to address infectious diseases, cancer, and future pandemics. Toll-like receptors (TLRs) have been researched as targets of vaccine adjuvants because they modulate the innate immune system and activate adaptive immunity. TLR-based adjuvants induce cross-presentation of exogenous antigens to CD8 T lymphocytes. TLR4 signaling recruits the adapters myeloid differentiation marker 88 (MyD88), which mediates the recycling of endoplasmic reticulum fusion to the phagosome as well as the TIR-domain-containing adapter-inducing interferon-β (TRIF), which is essential for translocation from the endosome to the cytosol.
Given the role of TLR4 agonists, most studies have used ligands based on lipopolysaccharides, but these have side effects (especially fever). Here we propose using TLR4 protein-based agonists (TLR-4-PBA), which have multiple advantages as adjuvants: They are naturally biocompatible and biodegradable substances that induce minimal reactogenicity and toxicity. We studied mollusk hemocyanin from Fissurella latimarginata (FLH) and the recombinant surface immunogenic protein from Streptococcus agalactiae (rSIP). These two proteins differ significantly in their structure, and we show Th1 immunomodulatory properties.
Using mouse bone marrow-derived dendritic cells pulsed with ovoalbumin (OVA), we then showed that rSIP and FLH promote cross-presentation in OT-I CD8+ T lymphocytes. This effect depends on TLR4 and the recruitment of MyD88 and TRIF. Studies are in progress to evaluate IRF3 and NF-κ B activation. Our results illustrate how these TLR-4-PBAs could act as a clinically useful vaccine adjuvant for innate immunity and Th1 adaptive immune response.
FONDECYT 1201600 (MIB); FONDEF ID21I10370 (AEV), Instituto de Salud Pública de Chile Funding (AEV, DD-D); ANID/PhD Nacional program 21210946 (MLS) and 21200880 (DD-D); FONDAP 15130011 (MT, SL).
The antigen presentation of MCMV specific Qa-1-restricted CD8+ T cells
Authors: Reilly Shanelle , Demers Delia , Fugere Celine , Brossay Laurent 
Published: August 2023

Abstract
Murine cytomegalovirus (MCMV) is a well-established model system for human CMV (HCMV), a ubiquitous herpesvirus that affects over 70% of people worldwide. Although the classical CD8+ T cell response to MCMV has been well characterized, little is known about the non-classical CD8+ T cell response. Our laboratory recently demonstrated that Qa-1-restricted CD8+ T cells are sufficient to protect mice from MCMV induced lethality. Our recent data shows that this protective T cell population has limited αβ TCR diversity. To identify the MCMV peptide(s) presented by Qa-1, we utilized bioinformatic tools that predict MHC binding, including NetMHC 4.0 and T Cell Epitope Prediction tools available on the Immune Epitope Database and Analysis Resource. Using these tools, we queried the entire MCMV coding sequence to identify potential nonamers predicted to bind to Qa-1. Furthermore, these candidates were also queried to identify protease cleavage sites to ensure that they are predicted to exist in vivo. Approximately 40 potential peptides were selected and tested in an in vitro screen using Qa-1 expressing cell lines. Our results suggest that several candidate peptides bind to Qa-1 as strongly as well-defined canonical binding partners. Taken together, these data suggest that an MCMV-responsive T cell population with limited αβ TCR diversity recognizes MCMV-derived peptides presented by Qa-1. This population is likely to be of importance for viral protection when classical CD8+ T cell immunity is compromised.
This work is supported by the NIH Research Grant AI046709. Shanelle Reilly is supported by research supplement R01 3AI046709-18S1 to promote diversity.
Mass spectrometric profiling of HLA-B44 peptidomes provides evidence for Tapasin-mediated Tryptophan editing
Authors: Kaur Amanpreet , Surnilla Avrokin , Zaitouna Anita J , Basrur Venkatesha , Mumphrey Michael B , Cieslik Marcin , Carrington Mary N , Nesvizhskii Alexey I , Raghavan Malini 
Published: August 2023

Abstract
Activation of CD8+ T cells against pathogens and cancers involves the recognition of antigenic peptides presented in complex with human leukocyte antigen (HLA) class-I proteins. Peptide binding to HLA-I proteins is coordinated by a multi-protein complex called the peptide loading complex (PLC). Tapasin, a key PLC component, facilitates the binding and optimization of HLA-I peptides. However, different HLA-I allotypes have variable requirements for tapasin for their assembly and surface expression. HLA-B*44:02 and HLA-B*44:05, which differ only at position 116 of their heavy chain sequences, fall at the opposite ends of the tapasin-dependency spectrum. HLA-B*44:02 (D116) is highly tapasin-dependent, whereas HLA-B*44:05 (Y116) is highly tapasin-independent. To better understand the influences of tapasin upon HLA-B44 peptidome compositions, we undertook mass spectrometric comparisons of HLA-B*44:02/05 peptidomes under tapasin-sufficient and/or tapasin-deficient conditions. Analyses of the HLA-B*44:05 peptidomes in the presence vs. absence of tapasin reveal that tapasin increases the frequency of C-terminal tryptophan residues. In the presence of tapasin, C-terminal tryptophans are also more highly represented among peptides unique to B*44:02 and those shared between B*44:02 and B*44:05, compared with peptides unique to B*44:05. Overall, our findings demonstrate that tapasin influences the C-terminal composition of HLA-I-bound peptides. For the HLA-B44 family, the presence of tapasin or high tapasin-dependence of an allotype results in better binding of peptides with C-terminal tryptophans, consistent with the role for tapasin in stabilizing an open conformation to accommodate bulky C-terminal residues.
Supported by grants from NIH to Malini Raghavan (R21 AI164025, R21 AI126054 and R01AI044115)
Expanded presentation of Lyme autoantigens and identification of a novel immunogenic CD4+ T cell epitope from Borrelia burgdorferi MCP4 in murine Lyme arthritis
Authors: Danner Rebecca , Pereckas Michael , Rouse Joseph , Wahhab Amanda , Lochhead Robert B 
Published: August 2023

Abstract
Lyme arthritis (LA), caused by Borrelia burgdorferi (Bb), is often accompanied by autoimmune T and B cell responses, but the mechanisms of infection-induced autoimmunity are unclear. We used an immunopeptidomics approach to identify potential Lyme autoantigens and immunogenic Bb antigens, which were validated by histology and testing of T cell reactivity. C57BL/6 (B6) mice, which develop mild inflammatory LA, and B6 Il10−/− (IL10 KO) mice, which develop severe, persistent LA, were infected with Bb for 4 or 16 weeks, and MHCII-bound peptides from inguinal and popliteal lymph nodes were identified by LC-MS/MS. Joint inflammation, fibrosis, and vascular remodeling were analyzed by H&E, Masson’s trichrome, and anti-CD31 immunohistochemistry, respectively. Six Bb peptides were identified by LC-MS/MS, of which one epitope from methyl-accepting chemotaxis protein 4 (MCP4) was an immunogenic CD4+ T cell antigen. Over 10,000 self peptides, particularly from proteins involved in cholesterol metabolism, tissue damage, and vascular inflammation were identified in infected mice. Presentation of peptides from previously identified human Lyme autoantigens apolipoprotein B-100, fibronectin, and type V collagen were expanded in infected mice, suggestive of epitope spreading. Consistent with immunopeptidomics data, joints showed increased inflammatory infiltrate, fibrosis and neovascularization in infected mice, compared with joints from uninfected mice. In conclusion, this immunopeptidomics approach revealed key insights into potential mechanisms of infection-induced autoimmunity in LA and indentified a novel immunogenic CD4+ T cell antigen from Bb MCP4.
Supported by grants from NIAID (R21AI148982)
QuickSwitch™: an MHC tetramer platform for making fast and reliable custom tetramers for T cell staining
Authors: Poluektov Yuri O , Delcommenne Marc C 
Published: August 2023

Abstract
The use of MHC tetramers to identify antigen specific T cells has become widespread among immunologists since the late 90s. Tetramers have become so ubiquitous that many companies and research facilities can ship and deliver some of the most commonly used tetramers within a matter of days. But when it comes to the uncommon tetramers or tetramers with antigenic peptides that have only recently been identified, it may be quite a challenge to find the needed tetramer. In this case, most researchers are left with trying to produce their own tetramer or having another facility do it for them, which can become a lengthy process. In an effort to help those researchers, we have developed the QuickSwitch™ MHC tetramer. This MHC tetramer comes loaded with a place-holder peptide (or exiting peptide) that can be exchanged for most well-binding peptides of a given MHC haplotype. This way, researchers are able to make a tetramer with any peptide they have available in a matter of hours. More importantly, our MHC tetramer platform comes with a means to monitor how much of the available peptide has exchanged with the exiting peptide and quantify the MHC occupancy. This allows the user to not only determine if the peptide in question can make a viable MHC tetramer, but also identify the strong and weak binding peptides for that MHC haplotype. This piece of information can be invaluable in future studies, in particular for vaccine development.
MBL International
Structures of antibody/MHC-I complexes reveal deficiencies in structure prediction programs
Authors: Boyd Lisa , Jiang Jiansheng , Ahmad Javeed , Natarajan Kannan , Margulies David H 
Published: August 2023

Abstract
The remarkable success of structural prediction programs such as AlphaFold2 and RosettaFold have revolutionized the contemporary approach to protein structure determination. However, intrinsically disordered proteins (IDPs) and protein/protein complexes provide a greater challenge to such algorithms. In particular, there has been limited success in the structural prediction of antibody/protein antigen complexes and TCR/MHC complexes, in part because the database of such structures is limited. Here, we present several high-resolution X-ray structures of anti-MHC/MHC-I complexes consistent with previous exon-shuffling and epitope mapping data that contradict computational predictions. Specifically, two mAb previously mapped to the alpha3 domain of H2-D(d) or H2-L(d)/D(b) respectively are shown to bind in distinct orientations, either of which may sterically block CD8 binding. In addition, a peptide dependent, but peptide non-specific anti-H2-D(d) mAb binds to loops of the alpha2 domain. These results substantiate the need to determine experimental structures of antibody/protein complexes as we proceed to develop avenues for the de novo design of specific antibodies. Supported by the Intramural Research Program of the NIAID, NIH
Supported by the Intramural Research Program of the NIAID, NIH
Endo-lysosomal assembly variations among Human Leukocyte Antigen class I (HLA-I) allotypes
Authors: Raghavan Malini , Olson Eli , Ceccarelli Theodora 
Published: August 2023

Abstract
The extreme polymorphisms of HLA-I proteins enable the presentation of diverse peptides to cytotoxic T lymphocytes (CTL). The canonical endoplasmic reticulum (ER) HLA-I assembly pathway enables presentation of cytosolic peptides, but effective intracellular surveillance requires multi-compartmental antigen sampling. Endo-lysosomes are generally sites of HLA class II assembly, but human monocytes and monocyte-derived dendritic cells (moDCs) also contain significant reserves of endo-lysosomal HLA-I molecules. We hypothesized variable influences of HLA-I polymorphisms upon outcomes of endo-lysosomal trafficking, as the stabilities and peptide occupancies of cell surface HLA-I are variable. Consistent with this model, when the endo-lysosomal pH of moDCs is disrupted, HLA-B allotypes display varying propensities for reductions in surface expression, with HLA-B* 08:01 or HLA-B*35:01 being among the most resistant or sensitive respectively, among eight tested HLA-B allotypes. Perturbations of moDC endo-lysosomal pH result in redistribution of HLA-B*35:01, but not HLA-B* 08:01, to LAMP1+ compartments and increase HLA-B*35:01 peptide receptivity. These findings reveal the intersection of the vacuolar cross-presentation pathway with a constitutive assembly pathway for some HLA-B allotypes. Notably, cross-presentation of epitopes derived from two soluble antigens was also more efficient for B*35:01 compared to B*08:01, even when matched for T cell response sensitivity, and more affected by cathepsin inhibition. Thus, HLA-I polymorphisms dictate the degree of endo-lysosomal assembly, which can supplement ER assembly for constitutive HLA-I expression and increase the efficiency of cross-presentation.
Supported by grants NIH R01 AI044115 T32GM008353 and T32AI007413
Inhibited MHC Class I and MHC Class II antigen processing and presentation upon Sars-CoV-2 infection
Authors: Nanaware Padma Pandurang , Cruz John , Khaja Muneeruddin , Shaffer Scott A , Stern Lawrence 
Published: August 2023

Abstract
Major histocompatibility (MHC) bound viral peptides are the immune signatures to initiate the selective and cross-reactive T cell responses. Identifying the antigenic targets of adaptive immunity to SARS-CoV-2 is a key towards understanding the pathogenesis of Sars-CoV-2 and the rapidly evolving mutants. We isolated the immunopeptidomes to identify naturally processed and presented MHC-II (HLA-DR, HLA-DP) and MHC-I (HLA-ABC) bound canonical and out-of-frame viral peptides. We identified 5 HLA-DR canonical epitopes (13 peptides) from spike protein, 2 HLA-DP canonical epitopes (4 peptides) from spike and membrane proteins, 1 canonical MHC-Class I peptides and 5 out-of-frame MHC-I peptides from different ORF proteins. The viral peptides were presented in lower abundances as compared to host peptides obscuring the identification of the MHC bound viral peptides. Some of these peptides were shown to recall T cell responses in COVID-19 donors in other reported studies. We observed the downregulation of surface and total HLA-ABC, HLA-DR and HLA-DP expression upon Sars-CoV-2 infection possibly restricting the presentation of viral peptides and therefore delaying the immune response. The mechanisms by which SARS-CoV-2 evades adaptive immune responses mediated by MHC-I are studied in detail by different groups. Hence, we further investigating the modulators of the MHC-II pathway using proteomics-based approach. We observed significant downregulation of CD74 protein, CIITA protein and cathepsins in infected cells affecting the processing and presentation of viral peptides. These results may help in understanding significantly altered antigen presentation in Sars-CoV-2 infected cells.
Structural classification of peptide/HLA complexes enables accurate high-throughput modeling
Authors: Gupta Sagar , Nerli Santrupti , Kandy Sreeja Kutti , Mersky Glenn L , Sgourakis Nikolaos G 
Published: August 2023

Abstract
The class I major histocompatibility complex (MHC-I) proteins display 8 to 15 residue peptides derived from endogenous proteins on the cell surface for immune surveillance. Accurate structural modeling of peptides bound to HLA (the human MHC) proteins has been mired by the inability to capture the central residues of the peptide. Here, an analysis of high-resolution X-ray crystal structures revealed that diverse peptide conformations arise as a result of local changes in the backbone configuration. As a result, we established an internal-coordinate based system to assess structural similarity between peptide backbones and found that peptide/HLA complexes, encompassing a wide range of allotypes and peptide sequences, present a discrete set of peptide backbone conformations. Leveraging these representative backbones and a regression model trained on a physically realistic scoring function, we developed RepPred, a structural modeling approach which predicts near native nonamer/HLA-A*02:01 complexes with high accuracy and throughput. Our method significantly outperformed five state-of-the-art peptide/HLA modeling approaches on the basis of structural accuracy. Thus, the structural information obtained from RepPred can be used to address long-standing immunology challenges such as predicting peptide immunogenicity.
Supported by grants from NIH (R01AI143997, R35GM125034, U01DK112217).
Nanoparticle geometry, immune memory, and antigen presentation determine the cross-reactive antibody response against sarbecoviruses
Authors: Wang Eric , Cohen Alexander A , Caldera Guzman Luis F , Bjorkman Pamela J , Chakraborty Arup K 
Published: August 2023

Abstract
In response to the threat of emerging SARS-CoV-2 variants and SARS-like betacoronaviruses (sarbecoviruses), mosaic nanoparticles presenting receptor-binding domains (RBDs) of multiple sarbecoviruses have been developed to elicit cross-reactive antibodies that target conserved regions of the RBD. To better understand vaccination with these antigens, we developed a computational model of the humoral response to nanoparticle and spike antigens using a combination of molecular dynamics, Markov processes, and population dynamics simulations. Avidity is currently only accounted for by introducing a second arm on rate, but additional effects of avidity will be considered in the future. Our model agrees with titers measured from nanoparticle and spike vaccinations in animals as well as clinical data from mRNA vaccines. We then used our model to predict the effect of previous vaccinations with wildtype spike on nanoparticle vaccinations. Compared to naïve individuals, previously vaccinated individuals produce a higher cross-reactive titer on the first nanoparticle vaccination and a lower cross-reactive titer on the second. This difference is due to expansion of cross-reactive B cells and competition with strain-specific B cells from previous vaccinations. Furthermore, we find that a cocktail of homotypic nanoparticles produces the same cross-reactive titers as the mosaic nanoparticle, which is because cross-reactive B cells can bind a greater fraction of presented antigen and can survive successive rounds of selection when the nanoparticle density is low.
Supported by grants from the National Science Foundation (1745302), NIH (1-R61- AI161805-01), and the Ragon Institute of MGH, MIT, and Harvard.