Sho Yamasaki

Mycobacteria have a cell envelope that can act as a shield against host defense. This study shows that mycobacteria survive in host macrophages by targeting the innate host receptor dectin-1 through a noncanonical ligand. Compared with wild-type (WT) mice, dectin-1–deficient mice were more resistant to infection to mycobacteria. Dectin-1–deficient mice presented with substantially reduced bacterial burdens, inflammatory cytokines, and infiltrating myeloid cells, such as neutrophils and macrophages. Intracellular survival of these bacteria was reduced in macrophages derived from dectin-1–deficient mice compared with those from WT mice. Cellular characterization of mycobacteria-infected macrophages indicated that the presence of dectin-1 altered phagosomal maturation and association with markers of autophagy. Activity-based purification and nuclear magnetic resonance spectrometry identified branched α-glucan as the dectin-1 mycobacterial ligand. This branched glucan was essential for activating dectin-1. These results show that mycobacterial α-glucan targets dectin-1 to facilitate intracellular bacterial survival.

MHC class I–related protein (MR1) presents vitamin B–based antigens (Ags) to mucosal-associated invariant T (MAIT) cells. While microbial riboflavin (RF) precursors are well-documented MR1 ligands, it is unclear whether host-generated RF catabolites influence MR1 immunity. Here, we report that RF catabolites, including 10-formylmethylflavin (FMF), lumichrome, lumiflavin, and alloxazine, bind to MR1 with moderate affinity, while RF itself binds weakly. In contrast to the MR1-upregulating microbial RF precursors, RF catabolites reduced the surface level of MR1 by inducing its retention in the endoplasmic reticulum and inhibiting exit. These RF catabolites weakly competed with vitamin B–based Ags for MR1 binding, thereby selectively inhibiting MAIT activation. The crystal structures of MR1 with RF, FMF, lumiflavin, and lumichrome show binding in the A′-pocket of MR1. Here, lumichrome formed a “flavin bond” covalent interaction with MR1-Lys43 differing from the typical Schiff base. Collectively, we identified three-ringed isoalloxazines that bind MR1 and reduce surface levels, suggesting a potential role in dampening MAIT cell immunity.

Development of invariant natural killer T (iNKT) cells in the thymus requires cell–cell interaction through invariant TCR (iTCR) and CD1d, which induces expression of the transcription factor, promyelocytic leukemia zinc finger (PLZF). However, the signaling pathway linking iTCR and PLZF remains unclear. Here, we report that a serine/threonine kinase, protein kinase D (PKD), plays a pivotal role in iNKT cell development. In T cell–specific PKD-deficient (Prkd2/3∆CD4) mice, PLZF induction and iNKT cell generation were severely impaired, which were rescued by introduction of a PLZF transgene. We identified the transcription factor Ikaros as a substrate of PKD upon iTCR stimulation. Knock-in mice carrying a phosphorylation-defective mutant Ikaros (Ikzf1S267/275A) exhibited an impairment of iNKT cell development, whereas conventional T cells were normal. In iNKT cells, Ikaros binds to the upstream region of the PLZF gene to induce its transcription. Mutant mice lacking the Ikaros-binding site (Zbtb16∆IBS) generated fewer iNKT cells than WT mice. These results suggest that PKD links iTCRs to PLZF induction through Ikaros, thereby mediating iNKT cell development.

Mincle, a member of the C-type lectin receptor (CLR) family, detects various glycolipids and glycerolipids, such as trehalose dimycolate (TDM) from Mycobacterium tuberculosis, leading to the activation of the innate immune system. In this study, we developed new fluorescence-labeled molecular probes, N-NBD-labeled TDE and O-NBD-labeled TDE, based on the structure of trehalose diester to elucidate the intracellular behavior of Mincle and its ligands. N-NBD-labeled TDE was prepared for the ligand analysis, and O-NBD-labeled TDE was specifically designed to label Mincle by Turn-ON fluorescent affinity labeling. Live-cell imaging analysis using these probes revealed that N-NBD-labeled TDE internalizes into the cell in a Mincle-dependent manner. Furthermore, imaging analysis using O-NBD-labeled TDE successfully detected Mincle in cells in a Mincle expression-dependent manner.

Our bodies are continuously exposed to injurious insults by infection and tissue damage, which are primarily sensed by innate immune receptors to maintain homeostasis. Among such receptors is macrophage-inducible C-type lectin (Mincle, gene symbol CLEC4E ), a member of the C-type lectin receptor (CLR) family, which functions as an immune sensor for both pathogens and damaged self. To monitor these injurious stimuli, Mincle recognizes disaccharide-based pathogen-derived glycolipids and monosaccharide-based intracellular metabolites, such as β-glucosylceramide. Mincle is well-conserved among mammals; however, there are questions that remain unclear, such as from which lower vertebrate did it arise and whether the original ligand was self or non-self. Here, we found homologues of Mincle and its signaling subunit Fc receptor γ chain (FcRγ) in lower vertebrates, such as reptiles, amphibians, and fishes. The crystal structure of a Mincle homologue revealed that fish Mincle possesses a narrower sugar-binding pocket than that of mammalian Mincle, and accommodates only monosaccharide moieties. These results suggest that Mincle may have evolved from a self-recognizing receptor, and its sugar-binding pocket widened during evolution, presumably to adapt to disaccharide-based glycolipids derived from life-threatening pathogens.

Background: The durability of vaccine-induced immune memory to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is crucial for preventing infection, especially severe disease.Methods: This follow-up report from a phase 1/2 study of S-268019-b (a recombinant spike protein vaccine) after homologous booster vaccination confirms its long-term safety, tolerability, and immunogenicity.Results: Booster vaccination with S-268019-b resulted in an enhancement of serum neutralizing antibody (NAb) titers and a broad range of viral neutralization. Single-cell immune profiling revealed persistent and mature antigen-specific memory B cells and T follicular helper cells, with increased B-cell receptor diversity. The expansion of B- and T-cell repertoires and presence of cross-reactive NAbs targeting conserved epitopes within the receptor-binding domain following a booster accounted for the broad-spectrum neutralizing activity.Conclusion: These findings highlight the potential of S-268019-b to provide broad and robust protection against a range of SARS-CoV-2 variants, addressing a critical challenge in the ongoing fight against coronavirus disease 2019 (COVID-19).