Biology subjects

Adams, E. J.

Publications and source records attributed to Adams, E. J..

8 recordsLinked to original sources

bioRxiv2026

Allosteric disulfide control of ligand binding and endocytosis of the natural killer cell receptor for HLA-G

Human Leukocyte Antigen (HLA)-G is selectively expressed by fetal trophoblast cells that invade maternal tissue and encounter maternal natural killer (NK) cells early in pregnancy. In NK cells, the endosomal receptor KIR2DL4 responds to soluble HLA-G by inducing a broad transcriptional program to support placental development. Structural features of KIR2DL4 that control ligand binding and endocytosis are unclear. Random mutagenesis revealed that three cysteines in the first immunoglobulin domain of KIR2DL4 regulate endocytosis and uptake of HLA-G. We found that the atypical Cys10-Cys28 disulfide bond observed in the KIR2DL4 crystal structure is an allosteric disulfide with potential to switch to a conventional Cys28-Cys74 bond. KIR2DL4 in human cells exists in both disulfide-bonded states, as shown by mass spectrometry analysis. The Cys10-Cys28 bond in a Cys74Ser KIR2DL4 mutant was reduced by protein disulfide isomerase (PDI) in vitro. Inhibition of PDI prevented HLA-G uptake by KIR2DL4 in both transfected 293T cells and primary NK cells. Mutants in the Cys10-Cys28 configuration endocytosed spontaneously but did not bind HLA-G. Conversely, KIR2DL4 with a Cys28-Cys74 bond was at the plasma membrane and responded to soluble HLA-G by endocytosis and transcription of the interferon stimulated gene IFI44L, like wild-type cells. A purified Cys10Leu KIR2DL4 mutant, which exhibited a reduced tendency to oligomerize, bound HLA-G in a peptide dependent manner with an affinity in the low micromolar range. Disulfide switching from the Cys10-Cys28 to the Cys28-Cys74 form correlated with a distant allosteric change, as predicted by AlphaFold, that could reorient a D0 domain loop to facilitate HLA-G binding. Thus, conversion from an inactive state to an HLA-G binding form regulates KIR2DL4 cellular localization and function to promote fetal development.

immunology↗

bioRxiv2024

Molecular characterization of the archaic HLA-B*73:01 allele reveals presentation of a unique peptidome and skewed engagement by KIR2DL2

HLA class I alleles of archaic origin may have been retained in modern humans because they provide immunity against diseases to which archaic humans had evolved resistance. According to this model, archaic introgressed alleles were somehow distinct from those that evolved in African populations. Here we show that HLA-B*73:01, a rare allotype with putative archaic origins, has a relatively rare peptide binding motif with an unusually long-tailed peptide length distribution. We also find that HLA-B*73:01 combines a restricted and unique peptidome with high-cell surface expression, characteristics that make it well-suited to combat one or a number of closely-related pathogens. Furthermore, a crystal structure of HLA-B*73:01 in complex with KIR2DL2 highlights differences from previously solved structures with HLA-C molecules. These molecular characteristics distinguish HLA-B*73:01 from other HLA class I alleles previously investigated and may have provided early modern human migrants that inherited this allele with a selective advantage as they colonized Europe and Asia.

biochemistry↗

bioRxiv2024

Disruption of riboflavin biosynthesis in mycobacteria establishes 5-amino-6-D-ribitylaminouracil (5-A-RU) as key precursor of MAIT cell agonists

Mucosal-associated invariant T (MAIT) cells exhibit an intrinsic ability to recognize and respond to microbial infections. The semi-invariant antigen recognition receptor of MAIT cells specifically detects the non-polymorphic antigen-presenting molecule, major histocompatibility complex class I-related protein 1 (MR1), which primarily binds riboflavin-derived metabolites of microbial origin. To further interrogate the dependence of these antigens on riboflavin biosynthesis in mycobacteria, we deleted individual genes in the riboflavin biosynthesis pathways in Mycobacterium smegmatis (Msm) and Mycobacterium tuberculosis (Mtb) and evaluated the impact thereof on MAIT cell activation. Blocking the early steps of the pathway by deletion of RibA2 or RibG profoundly reduced, but did not completely ablate, MAIT cell activation by Msm or Mtb, whereas deletion of RibC, which catalyzes the last step in the pathway, had no significant effect. Interestingly, deletion of RibH specifically enhanced MAIT cell recognition of Mtb whereas loss of lumazine synthase (RibH) activity had no impact on MAIT cell activation by Msm. MAIT cell activation by Msm was likewise unaffected by blocking the production of the MAIT cell antagonist, Fo (by inhibiting its conversion from the riboflavin pathway intermediate, 5-amino-6-D-ribitylaminouracil (5-A-RU), through the deletion of fbiC). Together, these results confirm a central role for 5-AR-U in generating mycobacterial MR1 ligands and reveal similarities and differences between Msm and Mtb in terms of the impact of riboflavin pathway disruption on MAIT cell activation. Author summaryMucosal-associated invariant T (MAIT) cells are an abundant population of innate-like T-cells which respond to microbial infections. These specialized cells recognize the MR1 molecule, which presents microbial metabolites derived from riboflavin (vitamin B2) biosynthesis. These cells are enriched in the airways and in some cases reduced in the peripheral blood of tuberculosis (TB) infected individuals suggestive of a role in the early response to infection by Mycobacterium tuberculosis. In this study, we investigated the effect of deleting individual genes in the riboflavin biosynthesis pathway on MAIT cell activation by Mycobacterium tuberculosis or Mycobacterium smegmatis. Our findings revealed that disrupting early stages in the pathway profoundly reduced but did not eliminate MAIT cell activation by both mycobacterial species. However, blocking the penultimate step in the pathway, catalyzed by the enzyme, lumazine synthase, led specifically to increased MAIT cell recognition of M. tuberculosis. Our results confirm the pivotal role of the riboflavin pathway intermediate, 5-A-RU, in generating mycobacterial ligands that serve as MAIT cell agonists. By enhancing our understanding of how MAIT cells recognize mycobacterial infections, the results of this study could inform strategies for the development of vaccines and/or immunotherapies for TB.

immunology↗

bioRxiv2023

Ligand-induced segregation from large cell-surface phosphatases is a critical step in γδ TCR triggering

Gamma/delta ({gamma}{delta}) T cells are unconventional adaptive lymphocytes that recognize structurally diverse ligands via somatically-recombined antigen receptors ({gamma}{delta} TCRs). The molecular mechanism by which ligand recognition initiates {gamma}{delta} TCR signaling, a process known as TCR triggering, remains elusive. Unlike {beta} TCRs, {gamma}{delta} TCRs are not mechanosensitive, and do not require coreceptors or typical binding-induced conformational changes for triggering. Here, we show that {gamma}{delta} TCR triggering by nonclassical MHC class Ib antigens, a major class of ligands recognized by {gamma}{delta} T cells, requires steric segregation of the large cell-surface phosphatases CD45 and CD148 from engaged TCRs at synaptic close contact zones. Increasing access of these inhibitory phosphatases to sites of TCR engagement, by elongating MHC class Ib ligands or truncating CD45/148 ectodomains, abrogates TCR triggering and T cell activation. Our results identify a critical step in {gamma}{delta} TCR triggering and provide insight into the core triggering mechanism of endogenous and synthetic tyrosine-phosphorylated immunoreceptors.

immunology↗

bioRxiv2022

Delivery of loaded MR1 Monomer Results in Efficient Ligand Exchange to Host MR1 and Subsequent MR1T cell activation

MR1-restricted T cells have been implicated in microbial infections, sterile inflammation, wound healing and cancer. Similar to other antigen presentation molecules, evidence supports multiple, complementary MR1 antigen presentation pathways. To investigate ligand exchange pathways for MR1, we used MR1 monomers and tetramers loaded with 5-(2-oxopropylideneamino)-6-d-ribitylaminouracil (5-OP-RU) to deliver the antigen. Using MR1-deficient cells reconstituted with wild-type MR1 or MR1 molecules that cannot bind 5-OP-RU, we show that presentation of monomer-delivered 5-OP-RU is dependent on cellular MR1 and requires the transfer of ligand from the soluble molecule onto MR1 expressed by the antigen presenting cell. This mode of antigen delivery strengthens the evidence for post-ER ligand exchange pathways for MR1, which could represent an important avenue by which MR1 acquires antigens derived from endocytosed pathogens.

immunology↗

bioRxiv2022

Deaza-modification of MR1 ligands modulates recognition by MR1-restricted T cells

MR1-restricted T (MR1T) cells recognize microbial small molecule metabolites presented on the MHC Class I-like molecule MR1 and have been implicated in early effector responses to microbial infection. As a result, there is considerable interest in identifying chemical properties of metabolite ligands that permit recognition by MR1T cells, for consideration in therapeutic or vaccine applications. Here, we made chemical modifications to known MR1 ligands to evaluate the effect on MR1T cell activation. Specifically, we modified 6,7-dimethyl-8-D-ribityllumazine (DMRL) to generate 6,7-dimethyl-8-D-ribityldeazalumazine (DZ), and then further derivatized DZ to determine the requirements for retaining MR1 surface stabilization and agonistic properties. Interestingly, the IFN-{gamma} response toward DZ varied widely across a panel of T cell receptor (TCR)-diverse MR1T cell clones; while one clone was agnostic toward the modification, most displayed either an enhancement or depletion of IFN-{gamma} production when compared with its response to DMRL. To gain insight into a putative mechanism behind this phenomenon, we used in silico molecular docking techniques for DMRL and its derivatives and performed molecular dynamics simulations of the complexes. In assessing the dynamics of each ligand in the MR1 pocket, we found that DMRL and DZ exhibit differential dynamics of both the ribityl moiety and the aromatic backbone, which may contribute to ligand recognition. Together, our results support an emerging hypothesis for flexibility in MR1:ligand-MR1T TCR interactions and enable further exploration of the relationship between MR1:ligand structures and MR1T cell recognition for downstream applications targeting MR1T cells.

immunology↗

bioRxiv2020

Biochemical Patterns of Antibody Polyreactivity Revealed Through a Bioinformatics-Based Analysis of CDR Loops

Antibodies are critical components of adaptive immunity, binding with high affinity to pathogenic epitopes. Antibodies undergo rigorous selection to achieve this high affinity, yet some maintain an additional basal level of low affinity, broad reactivity to diverse epitopes, a phenomenon termed "polyreactivity". While polyreactivity has been observed in antibodies isolated from various immunological niches, the biophysical properties that allow for promiscuity in a protein selected for high affinity binding to a single target remain unclear. Using a database of nearly 1,500 polyreactive and non-polyreactive antibody sequences, we created a bioinformatic pipeline to isolate key determinants of polyreactivity. These determinants, which include an increase in inter-loop crosstalk and a propensity for an "inoffensive" binding surface, are sufficient to generate a classifier able to identify polyreactive antibodies with over 75% accuracy. The framework from which this classifier was built is generalizable, and represents a powerful, automated pipeline for future immune repertoire analysis.

immunology↗

bioRxiv2017

The Function of the COPII Gene Paralogs SEC23A and SEC23B Are Interchangeable In Vivo

SEC23 is a core component of the coat protein-complex II (COPII)-coated vesicle, which mediates transport of secretory proteins from the endoplasmic reticulum (ER) to the Golgi1-3. Mammals express 2 paralogs for SEC23 (SEC23A and SEC23B). Though the SEC23 gene duplication dates back >500 million years, both SEC23s are ~85% identical at the amino acid sequence level. In humans, deficiency for SEC23A or SEC23B results in cranio-lenticulo-sutural dysplasia4 or congenital dyserythropoietic anemia type II (CDAII), respectively5. The disparate human syndromes and reports of secretory cargos with apparent paralog-specific dependence6,7, suggest unique functions for the two SEC23 paralogs. Here we show indistinguishable intracellular interactomes for human SEC23A and SEC23B, complementation of yeast SEC23 by both human and murine SEC23A/B paralogs, and the rescue of lethality resulting from Sec23b disruption in zebrafish by a Sec23a-expressing transgene. Finally, we demonstrate that the Sec23a coding sequence inserted into the endogenous murine Sec23b locus fully rescues the mortality and severe pancreatic phenotype previously reported with SEC23B-deficiency in the mouse8-10. Taken together, these data indicate that the disparate phenotypes of SEC23A and SEC23B deficiency likely result from evolutionary shifts in gene expression program rather than differences in protein function, a paradigm likely applicable to other sets of paralogous genes. These findings also suggest the potential for increased expression of SEC23A as a novel therapeutic approach to the treatment of CDAII, with potential relevance to other disorders due to mutations in paralogous genes.

evolutionary biology↗