Abstract
ABSTRACTToxin neutralizing antibodies represent the major mode of protective immunity against a number of toxin-mediated bacterial diseases, including anthrax; however, the cellular mechanisms that lead to optimal neutralizing antibody responses remain ill defined. Here we show that the cellular binding pathway of anthrax protective antigen (PA), the binding component of anthrax toxin, determines the toxin neutralizing antibody response to this antigen. PA, which binds cellular receptors and efficiently enters antigen-presenting cells by receptor-mediated endocytosis, was found to elicit robust anti-PA IgG and toxin neutralizing antibody responses. In contrast, a receptor binding-deficient mutant of PA, which does not bind receptors and only inefficiently enters antigen-presenting cells by macropinocytosis, elicited very poor antibody responses. A chimeric protein consisting of the receptor binding-deficient PA mutant tethered to the binding subunit of cholera toxin, which efficiently enters cells using the cholera toxin receptor rather than the PA receptor, elicited an anti-PA IgG antibody response similar to that elicited by wild-type PA; however, the chimeric protein elicited a poor toxin neutralizing antibody response. Taken together, our results demonstrate that the antigen capture pathway can dictate the magnitudes of the total IgG and toxin neutralizing antibody responses to PA as well as the ratio of the two responses.
Highlights
IMPORTANCE Neutralizing antibodies provide protection against a number of toxinmediated bacterial diseases by inhibiting toxin action
We were led to examine the role that receptor binding plays in the antibody response to protective antigen (PA), since in our previous work, we demonstrated that spontaneous deamidation of asparagine residues in PA is associated with a loss in the ability of the antigen to elicit toxin neutralizing antibodies [12], and as shown by others, deamidated PA exhibits reduced binding to cells [13]
To test the possibility that binding of PA to its receptor plays a role in its ability to elicit an antibody response, we utilized a receptor binding-deficient (RBD) form of PA that does not bind to its cellular receptors due to mutations (N682A and D683A) in its receptor-binding region [14, 15]
Summary
IMPORTANCE Neutralizing antibodies provide protection against a number of toxinmediated bacterial diseases by inhibiting toxin action. We have used protective antigen (PA), the binding component of anthrax toxin, as a model antigen to investigate immune mechanisms important for the induction of robust toxin neutralizing antibody responses. We found that the pathway used by antigenpresenting cells to capture PA dictates the robustness of the neutralizing antibody response to this antigen These results provide new insights into immune mechanisms that play an important role in the induction of toxin neutralizing antibody responses and may be useful in the design of new vaccines against toxin-mediated bacterial diseases. Vaccines against toxin-mediated bacterial diseases, such as diphtheria, tetanus, and anthrax, protect by eliciting robust toxin neutralizing antibody responses. This work demonstrates the importance of the PA receptor-specific binding pathway in eliciting a neutralizing antibody response to PA and demonstrates that antigen uptake pathways can dictate the robustness of the neutralizing antibody response
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