The goals are to explain, at the biochemical and cellular levels, the high immunogenicity of the Listeria monocytogenes (LM) pore-forming toxin listeriolysin O (LLO). LLO provides dominant antigens for both CD4+ and CD8+ T cell responses on multiple MHC haplotypes. The LLO molecule has two notable dual functions, first, it is a strongly stimulatory protein (i.e., it has an """"""""adjuvant"""""""" effect), inducing cytokine production and cell activation on the antigen presenting cells (APC). Second, LLO is an exceptional donor of antigens to the major histocompatibility class II molecules (MHC-II), eliciting strong T cell responses when given as a purified protein. Our hypothesis is that the strong avidity for cholesterol-rich membranes makes LLO a highly efficient immunogen. We want to take advantage of this feature to develop new knowledge on antigen processing and presentation biochemistry and to determine the relationship of LLO toxicity with its immunogenicity. We have chosen to focus on examining LLO as a purified protein, separate from the LM that normally expresses it. The benefits of this strategy are twofold: first, we can examine the immunogenicity of LLO in the absence of any other bacterial product that could change the biology of the APC, and, or induce an inflammatory effect;and second, we can take advantage of the known biochemistry of LLO. Examining the immunogenicity of LLO is important in order to: i) understand the natural infection and how LM antigens are presented;ii) explain LLO as an immunogen and a carrier molecule for ectopic antigens;and iii) provide us with unique clues on the basic mechanisms of antigen presentation.
In Aim 1, we describe the generation of an extensive series of LLO mutants that will be used to examine their immunogenicity both in vitro and in vivo. Mutagenesis of key amino acid residues on LLO will be informative in that it will allow us to probe the biochemical and biological features that makes LLO such a strong immunogen;such as binding to membranes, trafficking pathways, possible association with other molecules involved in presentation, capacity to activate the APC, etc. Of course, the information will be of interest to those using LLO as a vaccine vehicle and to those who study cholesterol dependent cytolysins. The various LLOs will be tested in the context of basic presentation assays and used in all subsequent aims.
In Aim 2, we describe approaches for determining the localization and catabolism of LLO inside APC. The subcellular localization of LLO will be followed by different approaches including developing fluorescently-tagged LLOs. We also plan to examine the relationship between the induction of cell death and autophagic processes by LLO and its immunogenicity.
In Aim 3, we examine the adjuvanticity of LLO in three contexts: i) activation of APC, ii) priming of naive T cells, and iii) generation of protective immunity to LM.

Public Health Relevance

We investigate immunity to the protein listeriolysin O, the major component that causes Listeria monocytogenes to be pathogenic. This protein is unique in its binding to cell surfaces and engenders an extremely powerful response. Studying the biochemical and cellular basis of such a protein will give us insights on how antigens are presented during microbial infection.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
Project #
Application #
Study Section
Immunity and Host Defense Study Section (IHD)
Program Officer
Mills, Melody
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Washington University
Schools of Medicine
Saint Louis
United States
Zip Code
Valderrama, Carolina; Clark, Amy; Urano, Fumihiko et al. (2017) Listeria monocytogenes induces an interferon-enhanced activation of the integrated stress response that is detrimental for resolution of infection in mice. Eur J Immunol 47:830-840
MacDuff, Donna A; Reese, Tiffany A; Kimmey, Jacqueline M et al. (2015) Phenotypic complementation of genetic immunodeficiency by chronic herpesvirus infection. Elife 4:
Lee, Sang Hun; Carrero, Javier A; Uppaluri, Ravindra et al. (2013) Identifying the initiating events of anti-Listeria responses using mice with conditional loss of IFN-? receptor subunit 1 (IFNGR1). J Immunol 191:4223-34
Carrero, Javier Antonio (2013) Confounding roles for type I interferons during bacterial and viral pathogenesis. Int Immunol 25:663-9
Carrero, Javier A; Vivanco-Cid, Hector; Unanue, Emil R (2012) Listeriolysin o is strongly immunogenic independently of its cytotoxic activity. PLoS One 7:e32310
Carrero, Javier A; Unanue, Emil R (2012) Mechanisms and immunological effects of apoptosis caused by Listeria monocytogenes. Adv Immunol 113:157-74
Unanue, Emil R; Carrero, Javier A (2012) Studies with Listeria monocytogenes lead the way. Adv Immunol 113:1-5
Edelson, Brian T; Bradstreet, Tara R; Hildner, Kai et al. (2011) CD8?(+) dendritic cells are an obligate cellular entry point for productive infection by Listeria monocytogenes. Immunity 35:236-48
Edelson, Brian T; Bradstreet, Tara R; KC, Wumesh et al. (2011) Batf3-dependent CD11b(low/-) peripheral dendritic cells are GM-CSF-independent and are not required for Th cell priming after subcutaneous immunization. PLoS One 6:e25660
Aoshi, Taiki; Carrero, Javier A; Konjufca, Vjollca et al. (2009) The cellular niche of Listeria monocytogenes infection changes rapidly in the spleen. Eur J Immunol 39:417-25

Showing the most recent 10 out of 15 publications