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Automated Measurement of Cryptococcal Species Polysaccharide Capsule and Cell Body
Published on: January 11, 2018
Effects of altered N-glycan structures of Cryptococcus neoformans mannoproteins, MP98 (Cda2) and MP84 (Cda3), on
Su-Bin Lee1, Catia Mota1, Eun Jung Thak1
1Department of Life Science, College of Natural Science, Chung-Ang University, Seoul, 156-756, South Korea.
Abstract:
Cryptococcus neoformans is an opportunistic human fungal pathogen causing lethal meningoencephalitis. It has several cell wall mannoproteins (MPs) identified as immunoreactive antigens. To investigate the structure and function of N-glycans assembled on cryptococcal cell wall MPs in host cell interactions, we purified MP98 (Cda2) and MP84 (Cda3) expressed in wild-type (WT) and N-glycosylation-defective alg3 mutant (alg3Δ) strains. HPLC and MALDI-TOF analysis of the MP proteins from the WT revealed protein-specific glycan structures with different extents of hypermannosylation and xylose/xylose phosphate addition. In alg3Δ, MP98 and MP84 had truncated core N-glycans, containing mostly five and seven mannoses (M5 and M7 forms), respectively. In vitro adhesion and uptake assays indicated that the altered core N-glycans did not affect adhesion affinities to host cells although the capacity to induce the immune response of bone-marrow derived dendritic cells (BMDCs) decreased. Intriguingly, the removal of all N-glycosylation sites on MP84 increased adhesion to host cells and enhanced the induction of cytokine secretion from BMDCs compared with that on MP84 carrying WT N-glycans. Therefore, the structure-dependent effects of N-glycans suggested their complex roles in modulating the interaction of MPs with host cells to avoid nonspecific adherence to host cells and host immune response hyperactivation.
Insights
Cryptococcus neoformans mannoproteins (MPs) N-glycan structures influence host immune responses. Altered N-glycans on MPs affect immune cell interactions, impacting fungal pathogenesis.
Area of Science:
- Mycology
- Immunology
- Glycobiology
Background:
- Cryptococcus neoformans is an opportunistic pathogen causing severe meningitis.
- Cell wall mannoproteins (MPs) are key antigens involved in host immune recognition.
- N-glycans on MPs play a role in host-pathogen interactions.
Purpose of the Study:
- To investigate the structure and function of N-glycans on C. neoformans MPs.
- To determine how N-glycans affect host cell adhesion, uptake, and immune response.
- To elucidate the role of N-glycan structure in C. neoformans pathogenesis.
Main Methods:
- Purification of MP98 and MP84 from wild-type and alg3Δ mutant strains.
- High-Performance Liquid Chromatography (HPLC) and MALDI-TOF analysis for glycan structure determination.
- In vitro adhesion, uptake, and bone-marrow derived dendritic cell (BMDC) assays.
Main Results:
- WT MPs exhibited hypermannosylation and xylose/xylose phosphate addition.
- N-glycosylation-defective alg3Δ mutant MPs had truncated N-glycans (M5 and M7 forms).
- Altered N-glycans reduced BMDC immune response induction but did not affect host cell adhesion.
- Removal of N-glycosylation sites on MP84 increased host cell adhesion and cytokine secretion from BMDCs.
Conclusions:
- N-glycan structure on C. neoformans MPs significantly modulates host cell interactions.
- N-glycans play a complex role in balancing host cell adherence and immune response.
- Understanding N-glycan function is crucial for developing strategies against cryptococcal meningitis.
