Bacterial infection disrupts established germinal center reactions through monocyte recruitment and impaired metabolic adaptation.

Consecutive exposures to different pathogens are highly prevalent and often alter the host immune response. However, it remains unknown how a secondary bacterial infection affects an ongoing adaptive immune response elicited against primary invading pathogens. We demonstrated that recruitment of Sca-1+ monocytes into lymphoid organs during Salmonella Typhimurium (STm) infection disrupted pre-existing germinal center (GC) reactions. GC responses induced by influenza, plasmodium, or commensals deteriorated following STm infection. GC disruption was independent of the direct bacterial interactions with B cells and instead was induced through recruitment of CCR2-dependent Sca-1+ monocytes into the lymphoid organs. GC collapse was associated with impaired cellular respiration and was dependent on TNFα and IFNγ, the latter of which was essential for Sca-1+ monocyte differentiation. Monocyte recruitment and GC disruption also occurred during LPS-supplemented vaccination and Listeria monocytogenes infection. Thus, systemic activation of the innate immune response upon severe bacterial infection is induced at the expense of antibody-mediated immunity.

Innate virus-sensing pathways in B cell systemic autoimmunity.

Although all multicellular organisms have germ line-encoded innate receptors to sense pathogen-associated molecular patterns, vertebrates also evolved adaptive immunity based on somatically generated antigen receptors on B and T cells. Because randomly generated antigen receptors may also react with self-antigens, tolerance checkpoints operate to limit but not completely prevent autoimmunity. These two systems are intricately linked, with innate immunity playing an instrumental role in the induction of adaptive antiviral immunity. In this work, we review how inborn errors of innate immunity can instigate B cell autoimmunity. Increased nucleic acid sensing, often resulting from defects in metabolizing pathways or retroelement control, can break B cell tolerance and converge into TLR7-, cGAS-STING-, or MAVS-dominant signaling pathways. The resulting syndromes span a spectrum that ranges from chilblain and systemic lupus to severe interferonopathies.

B cell class switch recombination is regulated by DYRK1A through MSH6 phosphorylation.

Protection from viral infections depends on immunoglobulin isotype switching, which endows antibodies with effector functions. Here, we find that the protein kinase DYRK1A is essential for B cell-mediated protection from viral infection and effective vaccination through regulation of class switch recombination (CSR). Dyrk1a-deficient B cells are impaired in CSR activity in vivo and in vitro. Phosphoproteomic screens and kinase-activity assays identify MSH6, a DNA mismatch repair protein, as a direct substrate for DYRK1A, and deletion of a single phosphorylation site impaired CSR. After CSR and germinal center (GC) seeding, DYRK1A is required for attenuation of B cell proliferation. These findings demonstrate DYRK1A-mediated biological mechanisms of B cell immune responses that may be used for therapeutic manipulation in antibody-mediated autoimmunity.

YTHDF2 suppresses the plasmablast genetic program and promotes germinal center formation.

Antibody-mediated immunity is initiated by B cell differentiation into multiple cell subsets, including plasmablast, memory, and germinal center (GC) cells. B cell differentiation trajectories are determined by transcription factors, yet very few mechanisms that specifically determine early B cell fates have been described. Here, we report a post-transcriptional mechanism that suppresses the plasmablast genetic program and promotes GC B cell fate commitment. Single-cell RNA-sequencing analysis reveals that antigen-specific B cell precursors at the pre-GC stage upregulate YTHDF2, which enhances the decay of methylated transcripts. Ythdf2-deficient B cells exhibit intact proliferation and activation, whereas differentiation into GC B cells is blocked. Mechanistically, B cells require YTHDF2 to attenuate the plasmablast genetic program during GC seeding, and transcripts of key plasmablast-regulating genes are methylated and bound by YTHDF2. Collectively, this study reveals how post-transcriptional suppression of gene expression directs appropriate B cell fate commitment during initiation of the adaptive immune response.

The germinal center reaction depends on RNA methylation and divergent functions of specific methyl readers.

Long-lasting immunity depends on the generation of protective antibodies through the germinal center (GC) reaction. N6-methyladenosine (m6A) modification of mRNAs by METTL3 activity modulates transcript lifetime primarily through the function of m6A readers; however, the physiological role of this molecular machinery in the GC remains unknown. Here, we show that m6A modifications by METTL3 are required for GC maintenance through the differential functions of m6A readers. Mettl3-deficient GC B cells exhibited reduced cell-cycle progression and decreased expression of proliferation- and oxidative phosphorylation-related genes. The m6A binder, IGF2BP3, was required for stabilization of Myc mRNA and expression of its target genes, whereas the m6A reader, YTHDF2, indirectly regulated the expression of the oxidative phosphorylation gene program. Our findings demonstrate how two independent gene networks that support critical GC functions are modulated by m6A through distinct mRNA binders.

Syk degradation restrains plasma cell formation and promotes zonal transitions in germinal centers.

Germinal centers (GCs) are sites at which B cells proliferate and mutate their antibody-encoding genes in the dark zone (DZ), followed by affinity-based selection in the light zone (LZ). B cell antigen receptor (BCR) signals induce Syk activation followed by rapid phosphatase-mediated desensitization; however, how degradation events regulate BCR functions in GCs is unclear. Here, we found that Syk degradation restrains plasma cell (PC) formation in GCs and promotes B cell LZ to DZ transition. Using a mouse model defective in Cbl-mediated Syk degradation, we demonstrate that this machinery attenuates BCR signaling intensity by mitigating the Kras/Erk and PI3K/Foxo1 pathways, and restricting the expression of PC transcription factors in GC B cells. Inhibition of Syk degradation perturbed gene expression, specifically in the LZ, and enhanced the generation of PCs without affecting B cell proliferation. These findings reveal how long-lasting attenuation of signal transduction by degradation events regulates cell fate within specialized microanatomical sites.

Antigen-delivery through invariant chain (CD74) boosts CD8 and CD4 T cell immunity.

Eradication of tumors by the immune system relies on the efficient activation of a T-cell response. For many years, the main focus of cancer immunotherapy has been on cytotoxic CD8 T-cell. However, stimulation of CD4 helper T cells is critical for the promotion and maintenance of immune memory, thus a good vaccine should evoke a two-dimensional T-cell response. The invariant chain (Ii) is required for the MHC class II heterodimer to be correctly guided through the cell, loaded with peptide, and expressed on the surface of antigen presenting cells (APC). We previously showed that by replacing the Ii CLIP peptide by an MHC-I cancer peptide, we could efficiently load MHC-I. This prompted us to test whether longer cancer peptides could be loaded on both MHC classes and whether such peptides could be accommodated in the CLIP region of Ii. We here present data showing that expanding the CLIP replacement size leads to T-cell activation. We demonstrate by using long peptides that APCs can present peptides from the same Ii molecule on both MHC-I and -II. In addition, we present evidence that antigen presentation after Ii-loading was superior to an ER-targeted minigene construct, suggesting that ER-localization was not sufficient to obtain efficient MHC-II loading. Finally, we verified that Ii-expressing dendritic cells could prime CD4+ and CD8+ T cells from a naïve population. Taken together our study demonstrates that CLIP peptide replaced Ii constructs fulfill some of the major requirements for an efficient vector for cancer vaccination.

Fap2 Mediates Fusobacterium nucleatum Colorectal Adenocarcinoma Enrichment by Binding to Tumor-Expressed Gal-GalNAc.

Fusobacterium nucleatum is associated with colorectal cancer and promotes colonic tumor formation in preclinical models. However, fusobacteria are core members of the human oral microbiome and less prevalent in the healthy gut, raising questions about how fusobacteria localize to CRC. We identify a host polysaccharide and fusobacterial lectin that explicates fusobacteria abundance in CRC. Gal-GalNAc, which is overexpressed in CRC, is recognized by fusobacterial Fap2, which functions as a Gal-GalNAc lectin. F. nucleatum binding to clinical adenocarcinomas correlates with Gal-GalNAc expression and is reduced upon O-glycanase treatment. Clinical fusobacteria strains naturally lacking Fap2 or inactivated Fap2 mutants show reduced binding to Gal-GalNAc-expressing CRC cells and established CRCs in mice. Additionally, intravenously injected F. nucleatum localizes to mouse tumor tissues in a Fap2-dependent manner, suggesting that fusobacteria use a hematogenous route to reach colon adenocarcinomas. Thus, targeting F. nucleatum Fap2 or host epithelial Gal-GalNAc may reduce fusobacteria potentiation of CRC.