Research Article

Natural polyreactive IgA antibodies coat the intestinal microbiota

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Science  20 Oct 2017:
Vol. 358, Issue 6361, eaan6619
DOI: 10.1126/science.aan6619

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Programmed recognition of microbiota

Increasingly, we recognize that the gut is a specialized organ for maintaining microbial symbioses alongside nutritional functions. The gut produces large quantities of immunoglobulin A (IgA), which adheres to the surface of gut microbes. Bunker et al. discovered that antibodies produced by naïve small intestinal plasma cells are recirculated and enriched within Peyer's patches, independently of exogenous antigen and T cell help. The resulting polyreactive IgAs are released into the gut lumen and bind to microbial surface glycans, thus innately recognizing the gut microbiota. Polyreactive IgAs appear to be a product of the coevolution of host and microbiota to maintain symbiotic homeostasis.

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Structured Abstract

INTRODUCTION

Immunoglobulin A (IgA) is the most abundant mammalian antibody isotype, constituting more than 80% of all antibody-secreting plasma cells at steady state. IgA is particularly prevalent at barrier surfaces such as the intestinal mucosa, where it forms a first line of defense in conjunction with innate mediators, including mucus and antimicrobial peptides. IgA is thought to coat and contain the resident commensal microbiota and provide protection against enteric pathogens. IgA responses occur under normal homeostatic conditions and involve both T cell–dependent and T cell–independent pathways of differentiation in mucosa-associated lymphoid tissues such as Peyer’s patches. However, despite its abundance, the specificity of homeostatic IgA has long remained elusive.

RATIONALE

To elucidate the specificity and origins of homeostatic IgA, we performed unbiased, large-scale cloning and characterization of monoclonal antibodies (mAbs) from single murine IgA plasma cells and other B cell populations of different origins. All antibodies were expressed recombinantly with an IgG1 isotype to compare their reactivity independent of their monomeric or multimeric nature.

RESULTS

Panels of single cell–derived mAbs were cloned from various B cell and IgA plasma cell populations, and their microbiota-reactivity was characterized by using a combination of bacterial flow cytometry and 16S ribosomal RNA (rRNA) sequencing. Additionally, mAbs were assayed by enzyme-linked immunosorbent assay (ELISA) for polyreactivity—a peculiar property of certain antibodies that facilitates binding to a variety of structurally diverse antigens. Several insights emerged from this characterization: (i) Microbiota-reactive and polyreactive antibodies arose naturally in all naïve B cell populations but were significantly enriched among IgA-secreting plasma cells. (ii) Microbiota-reactive and polyreactive antibodies from naïve B cells and IgA plasma cells showed similar patterns of binding to a broad, but defined, subset of microbiota. This binding included many members of Proteobacteria but largely excluded those of Bacteroidetes and Firmicutes, the predominant phyla in the colon. Interestingly, broadly neutralizing antibodies against influenza virus, which had previously been shown to be frequently polyreactive, were also commonly microbiota-reactive and displayed binding patterns that resembled IgAs. These patterns of microbiota-reactivity thus appear to be a general property of polyreactive antibodies. (iii) The microbiota-reactive and polyreactive IgA repertoire emerged via a mechanism that was largely independent of T cell help or somatic hypermutation. Instead, naturally microbiota-reactive and polyreactive recirculating naïve B cells were selected to become IgA plasma cells in Peyer’s patches. Although some antibodies subsequently acquired somatic mutations, these did not substantially alter their reactivity. (iv) Differentiation of microbiota-reactive and polyreactive IgAs occurred independent of microbiota or exogenous dietary antigen. Analysis of germ-free mice and germ-free mice fed an antigen-free diet demonstrated that microbiota-reactive and polyreactive IgA plasma cells arose naturally, even in the absence of exogenous antigens.

CONCLUSION

We conclude that homeostatic intestinal IgAs are natural polyreactive antibodies with innate specificity to microbiota. These data suggest that IgA antibodies, though derived from the adaptive immune system, possess innate-like recognition properties that may facilitate adaptation to the vast and dynamic array of exogenous microbiota and dietary antigens encountered at mucosal surfaces.

Large-scale analysis of mAbs reveals the specificity and origins of homeostatic intestinal IgA.

Panels of single cell–derived mAbs were cloned from murine IgA plasma cells and other B cell populations and characterized for microbiota-reactivity by bacterial flow cytometry and 16S rRNA sequencing or for polyreactivity against structurally diverse antigens, including DNA, insulin, lipopolysaccharide (LPS), flagellin, albumin, cardiolipin, and keyhole-limpet hemocyanin (KLH) by ELISA. This approach revealed that intestinal IgAs are natural polyreactive antibodies with innate specificity to microbiota. FSC, forward scatter.

Abstract

Large quantities of immunoglobulin A (IgA) are constitutively secreted by intestinal plasma cells to coat and contain the commensal microbiota, yet the specificity of these antibodies remains elusive. Here we profiled the reactivities of single murine IgA plasma cells by cloning and characterizing large numbers of monoclonal antibodies. IgAs were not specific to individual bacterial taxa but rather polyreactive, with broad reactivity to a diverse, but defined, subset of microbiota. These antibodies arose at low frequencies among naïve B cells and were selected into the IgA repertoire upon recirculation in Peyer’s patches. This selection process occurred independent of microbiota or dietary antigens. Furthermore, although some IgAs acquired somatic mutations, these did not substantially influence their reactivity. These findings reveal an endogenous mechanism driving homeostatic production of polyreactive IgAs with innate specificity to microbiota.

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