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Toward an integrated model of capsule regulation in Cryptococcus neoformans.

ABSTRACT: Cryptococcus neoformans is an opportunistic fungal pathogen that causes serious human disease in immunocompromised populations. Its polysaccharide capsule is a key virulence factor which is regulated in response to growth conditions, becoming enlarged in the context of infection. We used microarray analysis of cells stimulated to form capsule over a range of growth conditions to identify a transcriptional signature associated with capsule enlargement. The signature contains 880 genes, is enriched for genes encoding known capsule regulators, and includes many uncharacterized sequences. One uncharacterized sequence encodes a novel regulator of capsule and of fungal virulence. This factor is a homolog of the yeast protein Ada2, a member of the Spt-Ada-Gcn5 Acetyltransferase (SAGA) complex that regulates transcription of stress response genes via histone acetylation. Consistent with this homology, the C. neoformans null mutant exhibits reduced histone H3 lysine 9 acetylation. It is also defective in response to a variety of stress conditions, demonstrating phenotypes that overlap with, but are not identical to, those of other fungi with altered SAGA complexes. The mutant also exhibits significant defects in sexual development and virulence. To establish the role of Ada2 in the broader network of capsule regulation we performed RNA-Seq on strains lacking either Ada2 or one of two other capsule regulators: Cir1 and Nrg1. Analysis of the results suggested that Ada2 functions downstream of both Cir1 and Nrg1 via components of the high osmolarity glycerol (HOG) pathway. To identify direct targets of Ada2, we performed ChIP-Seq analysis of histone acetylation in the Ada2 null mutant. These studies supported the role of Ada2 in the direct regulation of capsule and mating responses and suggested that it may also play a direct role in regulating capsule-independent antiphagocytic virulence factors. These results validate our experimental approach to dissecting capsule regulation and provide multiple targets for future investigation.

SUBMITTER: Haynes BC

PROVIDER: S-EPMC3234223 | biostudies-literature | 2011 Dec

REPOSITORIES: biostudies-literature

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Toward an integrated model of capsule regulation in Cryptococcus neoformans.

Haynes Brian C BC Skowyra Michael L ML Spencer Sarah J SJ Gish Stacey R SR Williams Matthew M Held Elizabeth P EP Brent Michael R MR Doering Tamara L TL

PLoS pathogens 20111208 12

Cryptococcus neoformans is an opportunistic fungal pathogen that causes serious human disease in immunocompromised populations. Its polysaccharide capsule is a key virulence factor which is regulated in response to growth conditions, becoming enlarged in the context of infection. We used microarray analysis of cells stimulated to form capsule over a range of growth conditions to identify a transcriptional signature associated with capsule enlargement. The signature contains 880 genes, is enriche ...[more]

PMID: 22174677

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Project description:The polysaccharide capsule of Cryptococcus neoformans-an opportunistic basidiomycete pathogen and the major etiological agent of fungal meningoencephalitis-is a key virulence factor that prevents its phagocytosis by host innate immune cells. However, the complex signaling networks for their synthesis and attachment remain elusive. In this study, we systematically analyzed capsule biosynthesis and signaling networks using C. neoformans transcription factor (TF) and kinase mutant libraries under diverse capsule-inducing conditions. We found that deletion of GAT201, YAP1, BZP4, and ADA2 consistently caused capsule production defects in all tested media, indicating that they are capsule-regulating core TFs. Epistatic and expression analyses showed that Yap1 and Ada2 control Gat201 upstream, whereas Bzp4 and Gat201 independently regulate capsule production. Next, we searched for potential upstream kinases and found that mutants lacking PKA1, BUD32, POS5, IRE1, or CDC2801 showed reduced capsule production under all three capsule induction conditions, whereas mutants lacking HOG1 and IRK5 displayed enhanced capsule production. Pka1 and Irk5 controlled the induction of GAT201 and BZP4, respectively, under capsule induction conditions. Finally, we monitored the transcriptome profiles governed by Bzp4, Gat201, and Ada2 under capsule-inducing conditions and demonstrated that these TFs regulate redundant and unique sets of downstream target genes. Bzp4, Ada2, and Gat201 govern capsule formation in C. neoformans by regulating the expression of various capsule biosynthesis genes and chitin/chitosan synthesis genes in a positive and negative manner, respectively. In conclusion, this study provides further insights into the complex regulatory mechanisms of capsule production-related signaling pathways in C. neoformans. IMPORTANCE Over the past decades, human fungal pathogens, including C. neoformans, have emerged as a major public threat since the AIDS pandemic, only to gain more traction in connection to COVID-19. Polysaccharide capsules are rare fungal virulence factors that are critical for protecting C. neoformans from phagocytosis by macrophages. To date, more than 75 proteins involved in capsule synthesis and cell wall attachment have been reported in C. neoformans; however, their complex upstream signaling networks remain elusive. In this study, we demonstrated that Ada2, Yap1, Bzp4, and Gat201 were key capsule-inducing transcriptional regulators. Yap1 and Ada2 function upstream of Gat201, whereas Bzp4 and Gat201 function independently. Genome-wide transcriptome profiling revealed that Bzp4, Gat201, and Ada2 promote capsule production and attachment by positively and negatively regulating genes involved in capsule synthesis and chitin/chitosan synthesis, respectively. Thus, this study provides comprehensive insights into the complex capsule-regulating signaling pathway in C. neoformans.

Dataset Information

Toward an integrated model of capsule regulation in Cryptococcus neoformans.

Publications

Toward an integrated model of capsule regulation in Cryptococcus neoformans.

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