Allergen1 regulates polysaccharide structure in Cryptococcus neoformans

Abstract

Cryptococcus neoformans is an important human, fungal pathogen that sheds polysaccharide (exo-PS) into host tissues. While shed exo-PS mediates numerous untoward effects (including promoting increased intracranial pressure), little is known about the regulation of this phenomenon. Since down-regulation of the Allergen 1 (ALL1) gene is associated with high ICP, we investigated the relationship between ALL1 expression and exo-PS structure using a variety of biophysical techniques. The Δall1 mutants of two serotypes produced a shorter exo-PS with less branching and structural complexity than the parental strains. Consistent with lower branching, these exo-PSs manifested higher intrinsic viscosity than the parental strains. The Δall1 mutant strains manifested differences in epitope expression and significant resistance to phagocytosis. Exo-PS of Δall1 mutant exhibited anti-phagocytic properties. Comparative transcriptome analysis of mutant and parental strainunder iron-deprived conditions indicateda role of ALL1 in iron homeostasis, characterized by differential regulation of genes that mediate iron reduction and transport. Together, our results demonstrate a role of ALL1 in regulating conformational aspects of PS structure and iron homeostasis. These findings provide a mechanism to explain how changes in ALL1 expression influence virulence of switch variants and suggest that structural changes and polymer length are epigenetically regulated.

ALL2 , a Homologue of ALL1 , Has a Distinct Role in Regulating pH Homeostasis in the Pathogen Cryptococcus neoformans

“…Our results now demonstrate that the exo-PS of all2⌬ cells is augmented and has altered biophysical properties, namely, decreased viscosity compared to the exo-PS of the wild type. In contrast to this, the loss of ALL1 resulted in the secretion of a hyperviscous exo-PS and manifestation of a hypervirulent phenotype in murine models (12). The ALL2 mutant exhibits no significant growth defects in rich or nutrient-limiting media.…”

“…RNA transcript levels were measured by RT-PCR as previously described (7) in a 10-l reaction mixture containing SYBR green master mix (Applied Biosystems/Life Technologies) with the primers listed in Table S2 in the supplemental material. An RT-PCR analysis was performed with the appropriate controls, and the relative transcript levels were determined as described earlier (12).…”

“…Total RNA was isolated, and the microarray was performed by the Genome Technology Access Center at Washington University in St. Louis. Microarray analysis and gene ontology (GO) enrichment analysis were performed by the facility as described previously (12). Microarray results were further confirmed by performing RT-PCR on select genes as described above.…”

“…Under starvation, VAD1 downregulates ALL1 transcripts via degradation of mRNA (11). Although the precise function of ALL1 is not known, its expression indirectly affects polysaccharide conformation and iron homeostasis (12). Thus, intracisternal infection with the all1⌬ mutant results in augmented intracerebral pressure and premature death in infected rats (7).…”

Genome-Wide Transcription Study of Cryptococcus neoformans H99 Clinical Strain versus Environmental Strains

“…C . neoformans is an encapsulated yeast with sugar-coating, in which the sugar transport regulators are essential for the production of capsular polysaccharide antigens [ 34 ]. In fact, top in the list of the up-regulated genes in the H99 comprised SNF3 , a gene which is linked to the sugar transmembrane transport [ 35 , 36 ].…”