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Generation and characterization of a novel MHC-II tetramer for tracking and characterization of toxin B-specific CD4+ T cell responses
Journal article   Peer reviewed

Generation and characterization of a novel MHC-II tetramer for tracking and characterization of toxin B-specific CD4+ T cell responses

Jeffrey R Maslanka, Qianxuan She, Kathleen S Krauss, Emily N Bitsko, Nile U Bayard, Jennifer A Londregan, Mohamad-Gabriel Alameh, Laurence C Eisenlohr, Michele A Kutzler, Joseph P Zackular, …
The Journal of immunology (1950), v 215(8), vkag152
04 Aug 2026
DOI: https://doi.org/10.1093/jimmun/vkag152
PMID: 42601060
Featured in Collection :   Drexel's Newest Publications
url
https://doi.org/10.1093/jimmun/vkag152View
Published, Version of Record (VoR) Open

Abstract

Animals Bacterial Proteins - genetics Bacterial Proteins - immunology Bacterial Toxins - genetics Bacterial Toxins - immunology CD4-Positive T-Lymphocytes - immunology Clostridioides difficile - immunology Clostridium Infections - immunology Enterotoxins - immunology Epitopes, T-Lymphocyte - immunology Histocompatibility Antigens Class II - immunology Immunodominant Epitopes - immunology Mice Mice, Inbred C57BL
The gastrointestinal pathogen Clostridioides difficile is a major burden for health systems due to high rates of recurrence. C. difficile pathogenesis is mediated by two virulence factors, toxin A (TcdA) and toxin B (TcdB). Antibodies specific for TcdA and TcdB are correlated with protection from symptomatic recurrence; however, the role for CD4+ T cells is poorly understood in part due to the lack of tools to study the toxin-specific CD4+ T cell response. Our group recently demonstrated the antibody and CD4+ T cell response to C. difficile toxins is impaired via the glucosyltransferase activity of the toxins; however, tools do not exist to study the protective capacity and the phenotype of toxin-specific CD4+ T cells. Therefore, we developed a major histocompatibility complex class II (MHC-II) tetramer to identify TcdB-specific CD4+ T cells via flow cytometry. Herein, we identified an immunodominant epitope (TcdB1961-1975) in the CROPs region of TcdB and optimized an MHC-II tetramer for use in tracking and phenotyping TcdB-specific CD4+ T cell responses following multiple different immunization strategies in mice. Utilizing the tetramer, TcdB-specific T follicular helper cells were detected following TcdB-CROPs messenger RNA lipid nanoparticle vaccination validating the advantage of the tetramer. Furthermore, using a modular messenger RNA vector expressing the TcdB1961 peptide covalently bound to the beta chain of MHC-II, we were able to generate a robust population of TcdB-specific CD4+ T cells. These data outline the generation of new tools for the C. difficile field and lay the groundwork for future studies of toxin-specific CD4+ T cell responses.

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