Phagocytosis and Protein Processing Are Required for Presentation ofCryptococcus neoformansMitogen to T Lymphocytes

Abstract

In addition to eliciting antigen specific T-cell-mediated immunity, Cryptococcus neoformans possesses a mitogen (CnM) that activates naive T cells to proliferate. This mechanism of T-cell activation is accessory cell dependent and major histocompatibility complex unrestricted. CnM-induced T-cell proliferation correlates with internalization of the organism, suggesting that intracellular processing is required to liberate CnM prior to presentation to T cells. To determine whether phagocytosis and processing are required, various inhibitors of accessory cell uptake and processing were used. C. neoformans was observed within the accessory cells. Paraformaldehyde fixation of the accessory cell abrogated presentation of CnM to T cells, indicating that a dynamic accessory cell surface was required. A lysosomotropic agent abrogated the response to CnM but had no effect on a control stimulus that did not require processing. Both aspartic acid and cysteine protease inhibitors blocked effective processing of CnM, so that it was unable to stimulate T cells. Finally, an inhibitor of microfilament polymerization abrogated proliferation to CnM. These results indicate that the mitogenic activity of C. neoformans requires phagocytosis of the organism, lysosomal or endosomal processing, proteolytic activity, and microfilament polymerization and intracellular transport as a prerequisite for T-cell proliferation.Cryptococcus neoformans is a pathogenic yeast that causes one of the leading fatal mycoses in AIDS (9,12,18,38). This clinical observation, in addition to animal studies, has made it clear that T-cell-mediated immunity is of paramount importance in host defense against C. neoformans (1,7,15,19,20,22,28,31). T-cell immunity is antigen specific; however, we have recently shown that T cells are also capable of responding to C. neoformans by an alternate mechanism of activation (36). Specifically, when T cells from a previously unexposed individual are placed in culture with C. neoformans, the organism induces a mitogenic response. The mitogenic T-cell response is accessory cell (AC) dependent but not major histocompatibility complex restricted (36). Since the whole organism is capable of mediating this mitogenic effect, it raises questions about how the C. neoformans mitogen (CnM) might be liberated or displayed prior to T-cell activation.We had previously shown that CnM was confined to the cell wall of the organism (32). Since CnM may be displayed on the cell wall, it is possible that it cross-links surface molecules on the T cell and AC analogous to superantigens. Alternately, the AC might produce an enzyme that results in extracellular degradation and release of CnM, with liberation of the molecule into the surrounding milieu, where it exerts its mitogenic effect. Finally, we considered the possibility that the organism must be taken up by the AC and degraded internally, with subsequent presentation of CnM by the AC to the T cell. We have previously shown that lymphocyte proliferation in response to C. neoformans correlate...

Primary Dendritic Cells PhagocytoseCryptococcus neoformansvia Mannose Receptors and Fcγ Receptor II for Presentation to T Lymphocytes

“…It had been previously demonstrated that the lysosomal compartment and serine proteases were required for processing of C. neoformans (74). This raised the possibility that a leucine methyl ester (LME)-sensitive compartment might be required for processing of C. neoformans.…”

“…It was previously demonstrated that APC are required for T-cell responses to CnM (75) and that phagocytosis of C. neoformans is required for the APC to present CnM to T cells (73,74). To establish the type of cell that is the APC, blood mononuclear cells were separated based on their adherence properties, which have been used by other investigators to separate populations of APC (35,57,69).…”

“…The evidence for a C. neoformans mitogen (CnM) comes from the ability of CnM to stimulate the proliferation of naive T cells, the ability of allogeneic accessory cells to provide costimulatory activity, and the precursor frequency of responding T cells (49,52). Similar to some bacterial mitogens (2,41) and cognate peptide antigens (20), uptake and processing of C. neoformans by antigen-presenting cells (APC) was required prior to presentation of CnM to T cells (74).…”

Phenotypic switching of Cryptococcus neoformans occurs in vivo and influences the outcome of infection

“…Successful phagocytosis is required for effective CMI, and proliferation of lymphocytes in response to C. neoformans correlates with the magnitude of phagocytosis (46,47). Therefore, we hypothesized that impaired phagocytosis affects the host-pathogen interaction and alters the CMI.…”

Melanization ofCryptococcus neoformansAffects Lung Inflammatory Responses during Cryptococcal Infection

“…Hence, increased melanin activation of the complement system may have been responsible for the significantly enhanced opsonization of melanized nonencapsulated cells relative to that of nonmelanized nonencapsulated cells. Differences in the efficiency of phagocytosis of melanized and nonmelanized cells could translate into differences in the immune response since fungal cell ingestion can affect antigen presentation, cytokine production, dendritic cell activation, intracellular fungal growth, and macrophage cell viability (4,(34)(35)(36)(37).…”