Epitope Location in the Cryptococcus neoformans Capsule Is a Determinant of Antibody Efficacy

Abstract

Monoclonal antibodies (mAbs) to the polysaccharide capsule of Cryptococcus neoformans can prolong survival in mice. However, the properties of antibodies that mediate protection are not fully understood. The IgM mAbs 12A1 and 13F1 originated from the same B cell and differ only by somatic mutations in their variable regions; yet mAb 12A1 protects against serotype D infection, while mAb 13F1 does not. Phage peptide display libraries were used to analyze the fine specificity of these two mAbs. The selection of distinct peptide motifs from identical libraries confirmed that mAbs 12A1 and 13F1 bound to two distinct epitopes. Immunofluorescence and immunoelectron microscopy studies revealed differences in antibody localization within the capsule of serotype D strain; mAb 12A1 bound to the outer rim of the capsule resulting in an annular pattern, whereas mAb 13F1 bound throughout the capsule and had a punctate appearance. The difference in the binding pattern of mAb 12A1 and 13F1 was not observed on serotype A organisms, where both mAbs bound to the capsule with an annular fluorescence pattern. The fluorescence pattern of binding correlated with protective efficacy; mAb 13F1 prolonged survival of mice infected with the J11 serotype A strain (annular fluorescence), but not serotype D strains (punctate pattern). Annular binding, but not punctate binding, was associated with increased opsonic efficacy for phagocytosis of C. neoformans by J774.16 macrophage-like cells. The correlation between capsular binding pattern, opsonic activity, and ability to prolong survival suggests that the efficacy of anticryptococcal antibodies is dependent upon where they bind in the polysaccharide capsule.

Immunoglobulin M Efficacy AgainstCryptococcus neoformans: Mechanism, Dose Dependence, and Prozone-Like Effects in Passive Protection Experiments

“…Both mAbs activate the classical complement pathway, but mAb 12A1 suppresses capsular activation of the alternative complement pathway (16). The differences in immunofluorescence-binding pattern, capsular reaction, competition studies, and peptide mimotope reactivity indicate that these two mAbs bind to different epitopes in GXM (12,13,17).…”

“…These two IgMs originated from the same B cell, use the same V region, but differ in certain complementaritydetermining region amino acid sequences as a result of somatic mutations, which in turn result in differences in specificity (12). mAbs 12A1 and 13F1 both protect against infection with serotype A isolates, but only mAb 12A1 protects against serotype D infection (13). mAbs 12A1 and 13F1 produce annular and punctate fluorescence patterns with serotype D isolates, respectively (14).…”

“…The IgM mAbs 12A1 and 13F1 have been described previously (12,13,23). IgM ascites was generated in BALB/c mice given i.p.…”

B‐Cell Epitopes in the Immunodominant p34 Antigen of Mycobacterium avium ssp. paratuberculosis Recognized by Antibodies from Infected Cattle

“…For example, only some epitopes elicited a protective response in mice infected with C. neoformans [23]. Moreover, the protective response was also dependent of the gamma sub-isotype of murine antibodies used in passive transfer experiments [24].…”

The protective role of immunoglobulins in fungal infections and inflammation

“…In addition, mouse background also shown to influence the protection given by IgG subclass antibodies [68] thus underscoring the complex relationship between the cellular and humoral components of the immune system. Further studies from the same group revealed that epitope specificity of the MAbs is the critical factor that determines the serotype-specific protection rendered by the anti-C. neoformans MAbs and to confer protection against distinct serotypes of C. neoformans [69].…”