IgA Clonal Lineage Analysis Reveals Class Switch Dynamics in Human Gut

Introduction: IgA is the dominant antibody class in the gut, secreted by more than 75,000 IgA-producing plasma cells daily. Human IgA subtypes show distinct anatomical expression patterns, with IgA1 dominating in serum and IgA2 is abundant at sites colonized by a large microbiota, including the distal intestinal tract. IgA1 has a longer hinge region than IgA2, making it more vulnerable to degradation by bacterial proteases that target its hinge region. Direct IgM to IgA2 switching, rather than sequential switching through IgA1, may allow B cells to acquire resistance to BCR degradation. We aimed to estimate the relative frequencies of these two pathways.

Materials and methods: Ig RNA was obtained from cells sampled at the normal terminal ileum and the ascending colon of four patients undergoing right hemicolectomy and sequenced. Sequences were pre-processed using pRESTO (Vander Heiden et al., 2014), assigned for sub-isotype using a custom python script and annotated using IMGT/HighV-QUEST (Brochet, Lefranc, & Giudicelli, 2008). Clones were assigned using Change-O (Gupta et al., 2015) and sampling depth was assessed using rarefaction plots. Lineage trees were constructed using IgTree© (Barak, Zuckerman, Edelman, Unger, & Mehr, 2008) and analyzed using PopTree© and custom R scripts.

Results and discussion: Sampling depth of all the samples was good. More clones were shared between IgA1 and IgA2 than between IgM and IgA2. Lineage tree analysis revealed that the most abundant class switching in the ileum and colon is from IgA1 to IgA2, and that IgM cells tend to change location or population, rather than switching. Finally, direct IgM to IgA2 switching was significantly more abundant than IgM to IgA1; however, IgA1 to IgA2 switching was significantly much more abundant than IgM to IgA2.

Conclusion: Overall, these results support the idea that the majority of the IgA population in the intestine is created early in life. The IgM tendency to switch to IgA2 rather than IgA1, may allow B cells to acquire resistance to BCR degradation by bacterial proteases that target the hinge region of IgA (Cerutti, 2008).