B cells, the immune cells that produce antibodies, each recognize and bind a specific antigen through their B cell receptor (BCR). Genes within the BCR undergo random recombination during development, giving rise to B cell repertoires with immense diversity. However, the stochastic nature of B cell development also produces B cells specific for self-antigen which can cause serious autoimmune conditions. To eliminate auto-immune reactivity, several levels of checkpoints throughout B cell development prevent self-reactive B cells from developing or becoming activated. However, this system can also backfire, as some B cells specific for a foreign antigen may also display low-level cross-reactivity for self-antigen, leading to their deletion or hindrance. To “better understand B cell responses to vaccines,” the Taylor lab in the Vaccine and Infectious Disease Division studied “the diverse populations of naïve B cells targeting antigens relevant for public health as well as model antigens that are more easily studied,” Dr. Blair DeBuysscher explained. Dr. DeBuysscher, along with Holly Steach, a former Taylor lab technician who is now a PhD candidate at Yale, headed this study and recently published their work in the Journal of Immunology.
To determine to what degree naïve B cells specific for foreign antigen were cross-reactive to self-antigen, the authors used Nur77eGFP reporter mice whose B cells fluoresce following BCR stimulation in a signal strength-dependent manner. Because these B cells have never encountered the antigen for which they are specific, any Nur77eGFP expression, and therefore BCR stimulation, can be attributed to cross-reactivity to a self-antigen. From these mice, B cells specific for a “model” antigen and assessed for Nur77eGFP expression. The degree of cross-reactivity with self-antigen varied between B cells specific for various model antigens, but was not increased specifically in model antigen-specific B cells compared to polyclonal B cells of unknown specificities. Because model antigens are convenient tools that may not be representative of antigens derived from human pathogens, the authors next repeated this experiment with naïve B cells specific for HIV, respiratory syncytial virus (RSV) and influenza virus, and again found the same level of cross-reactivity to self-antigen. Together, these experiments showed that a range of cross-reactivity to self is observed within B cells of a given specificity, but that cross reactivity to self is not limited to B cells specific for foreign antigens.
Previous work demonstrates that cross-reactive B cells are less responsive when exposed to their cognate foreign antigen. To determine if this lack of responsiveness corresponded to a reduced ability to proliferate after BCR stimulation, Nur77eGFP-low, medium, and high B cells were identified as a proxy for increasing levels of cross-reactivity with self-antigen and incubated with anti-immunoglobulin to stimulate the BCR. Proliferation was measured by a cell dye that dilutes as cells divide, reavealing that Nur77eGFP-high B cells proliferated least in response to BCR signaling, while low Nur77eGFP-expressing B cells proliferated the most. However, these differences were lost when immunoglobulin concentrations were increased, suggesting that strong BCR signal can overcome the dampening effects of cross-reactivity to self. Based on these in vitro results, the authors moved to a mouse model to determine if cells displaying high self-reactivity would have dampened antigen responses in vivo. They transferred Nur77eGFP -low, med, and high dye-labeled cells into recipients immunized with their cognate model antigen. They found that Nur77eGFP-low and high cells proliferated least compared to Nur77eGFP-medium donor cells, suggesting that self-reactivity acts as a secondary signal to tune B cells responses to antigen and that a moderate level of self-reactivity corresponds with optimal B cell function.
Finally, to corroborate their results in humans, the authors identified HIV- and RSV-specific human B that expressed IgM, a type of BCR common in self-reactive B cells. As in mouse cells, human B cells specific for foreign antigens did not express differential amounts of IgM compared to bulk B cells, suggesting that cross-reactivity to self is a B cell-wide trait not limited to B cells of certain specificities. Likewise, the authors directly cloned HIV-specific BCR sequences and assessed binding to a human cell line, but found that HIV-specific B cells were no more self-reactive than naïve, polyclonal B cells.
Unlike what has been previously hypothesized, this study suggests that B cell cross-reactivity to self is not limited to B cells is a shared trait within all polyclonal populations of B cells targeting foreign antigens. The data suggests a model in which a B cell must have strong affinity for a foreign antigen to become activated, but that additional affinity for self-antigen further tunes B cell functionality. Dr. DeBuysscher further summarized their findings. “We found that both mouse and human antigen-specific naïve B cell populations contained cells that were also able to target self-antigens. By splitting the population into cells with high, moderate, and low reactivity to these self-antigens groups, we were able to pinpoint the effects of self-reactivity on B cell response after antigen stimulation.” Moving forward, these results have important implications for vaccine development. “Our data showed that moderate levels of self-reactivity lead to the optimal B cell responses when we vaccinated mice. This work suggests that we can predict which B cells are more likely to respond to candidate vaccines,” DeBuysscher said.
This work was supported by the National Institutes of Health.
UW/Fred Hutch Cancer Consortium member Justin Taylor contributed to this work.
Steach HR, DeBuysscher BL, Schwartz A, Boonyaratanakornkit J, Baker ML, Tooley MR, Pease NA, Taylor. 2020. Cross-Reactivity with Self-Antigen Tunes the Functional Potential of Naive B Cells Specific for Foreign Antigens. Journal of Immunology. 2020 Feb 1;204(3):498-509. doi: 10.4049/jimmunol.1900799. Epub 2019 Dec 27.