Binding of purified and radioiodinated capsular polysaccharides from Cryptococcus neoformans serotype A strains to capsule-free mutants

Abstract

Strains 6, 15, 98, 110, and 145 of Cryptococcus neoformans serotype A vary in capsule size, animal virulence, and susceptibility to in vitro phagocytosis. The isolated capsular polysaccharides (CPSs) differ in monosaccharide composition ratios and molecular size, as determined by gel filtration. The purpose of this investigation was to characterize the binding of CPSs to capsule-free mutants of C. neoformans and to examine CPSs from these strains for differences in their ability to bind, to determine whether such differences might explain the variation in the pathobiology of these strains. CPSs were partially periodate oxidized, tyraminated, iodinated with 1251, and used in binding studies with two capsule-free mutants of C. neoformans, strain 602 and Cap59. Binding was specific for yeast species and for polysaccharide and was saturable, which is consistent with a receptor-mediated mechanism of attachment. Binding occurred rapidly and was only slowly reversible. Binding was also independent of pH from pH 5.5 to 8, of cation concentrations, and of competition by sugars up to 1.0 M concentrations. Only a portion of CPS was capable of binding, and strains varied in the extent to which their CPS bound. CPS-15-IV (peak IV was the major polysaccharide peak on DEAE-cellulose chromatography of CPS from strain 15) had the highest proportion of binding (40%), followed by CPS from strains 98, 6, 145, 110, and 15-III (peak III was an earlier eluting fraction of CPS from strain 15). The CPSs differed similarly in their ability to competitively inhibit binding. Treatment of CPS, but not yeast cells, with proteinase XIV abolished binding without altering the CPS gross structure. Treatment of yeast cells with proteases, heat, or formaldehyde did not alter binding, and both strain 602 and Cap59 bound CPS similarly. Binding to encapsulated yeast cells was minimal.

Encapsulation ofCryptococcus neoformanswith Glucuronoxylomannan Inhibits the Antigen-Presenting Capacity of Monocytes

“…The observation that acapsular cryptococci can be incubated with purified GXM and washed and acquire a diminished ability to stimulate lymphoproliferation (Table 1) identifies GXM as the capsular component which mediates the suppression. These results confirm similar observations by ourselves (39,40,43) and others (5,26). A role for GXM in the suppressive phenomenon is further supported by our more recent study which found that anti-GXM MAbs are able to upregulate APC function of monocytes treated with encapsulated cryptococci (44).…”

“…Incubation of acapsular cryptococci with purified GXM leads to binding of GXM to the yeast surface (14,16,40) and…”

Downregulation by cryptococcal polysaccharide of tumor necrosis factor alpha and interleukin-1 beta secretion from human monocytes

“…If it is assumed that IL-1␤ secretion is a response to phagocytosis of the acapsular yeast cells, there is a strong possibility that GXM acts by inhibiting phagocytosis of the yeast. Addition of purified GXM to acapsular cryptococci leads to binding of polysaccharide to the cells (16,34), with a consequent inhibition of phagocytosis (3,12).…”

Spatial and temporal sequence of capsule construction in Cryptococcus neoformans

“…Experiments that demonstrate high-affinity binding of shed capsule polysaccharides to acapsular cells suggest that a saturable binding process may occur (Kozel and Hermerath, 1984;Small and Mitchell, 1986), but details of this association have not been established. Displacement of pre-existing capsule may contribute to the observed constitutive shedding of capsule into the surrounding environment, whether that is culture medium or the tissues and body fluids of an infected animal.…”

“…Alternatively, capsule components shed or secreted from the cell could diffuse outwards and associate with distal parts of the structure. This model (II) is suggested by experiments showing that capsular polysaccharide purified from culture medium can bind to acapsular mutants of Cryptococcus (Kozel, 1977;Kozel and Hermerath, 1984;Small and Mitchell, 1986). Lastly, new material may be interspersed with old (model III), or a combination of these scenarios might occur.…”