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Role of actin-bundling protein Sac6 in growth of Cryptococcus neoformans at low oxygen concentration.

ABSTRACT: Cryptococcus neoformans, the etiologic agent of cryptococcosis, is an obligately aerobic yeast that inhabits an environmental niche exposed to ambient air. The cell doubling time was significantly prolonged under 1% O(2) relative to that under normoxic conditions. No apparent cell cycle arrest occurred following a shift from ambient air to 1% O(2). However, yeast cells became hypersensitive to the actin monomer-sequestering agent latrunculin A at 1% O(2), indicating that proper actin function is critical for growth at low oxygen concentrations. We showed that Sac6, an actin-binding protein, played an important role in cell growth under low oxygen conditions. Sac6 colocalized with cortical actin patches and with the ring structures between mother cells and buds. Under low oxygen conditions, the sac6 deletion mutant grew poorly, and accumulation of the actin capping protein Cap1 was observed in the vacuole of the sac6? strain. Furthermore, endocytic processes were hampered in the sac6? mutant, but cell polarity and cytokinesis were not visibly disturbed. The deficiency of endocytosis in the sac6? strain could be rescued by 1 M sorbitol under 1% O(2), but growth remained retarded. These results suggest an absence of a direct link in C. neoformans between endocytosis and coping with the stress of low oxygen conditions. This interpretation is further supported by the observation that deletion of three conserved genes, ABP1, CRN1, and SLA2, which play important roles in endocytosis, had no effect on growth under 1% O(2). Interestingly, deletion of SAC6 in C. neoformans had no effect on virulence in mice.

SUBMITTER: Chang YC

PROVIDER: S-EPMC3416496 | biostudies-literature | 2012 Jul

REPOSITORIES: biostudies-literature

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Role of actin-bundling protein Sac6 in growth of Cryptococcus neoformans at low oxygen concentration.

Chang Yun C YC Lamichhane Ami Khanal AK Kwon-Chung Kyung J KJ

Eukaryotic cell 20120504 7

Cryptococcus neoformans, the etiologic agent of cryptococcosis, is an obligately aerobic yeast that inhabits an environmental niche exposed to ambient air. The cell doubling time was significantly prolonged under 1% O(2) relative to that under normoxic conditions. No apparent cell cycle arrest occurred following a shift from ambient air to 1% O(2). However, yeast cells became hypersensitive to the actin monomer-sequestering agent latrunculin A at 1% O(2), indicating that proper actin function is ...[more]

PMID: 22562467

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Project description:UNLABELLED:While research has identified an important contribution for metals, such as iron, in microbial pathogenesis, the roles of other transition metals, such as copper, remain mostly unknown. Recent evidence points to a requirement for copper homeostasis in the virulence of Cryptococcus neoformans based on a role for a CUF1 copper regulatory factor in mouse models and in a human patient cohort. C. neoformans is an important fungal pathogen that results in an estimated 600,000 AIDS-related deaths yearly. In the present studies, we found that a C. neoformans mutant lacking the CUF1-dependent copper transporter, CTR4, grows normally in rich medium at 37°C but has reduced survival in macrophages and attenuated virulence in a mouse model. This reduced survival and virulence were traced to a growth defect under nutrient-restricted conditions. Expression studies using a full-length CTR4-fluorescent fusion reporter construct demonstrated robust expression in macrophages, brain, and lung, the latter shown by ex vivo fluorescent imaging. Inductively coupled mass spectroscopy (ICP-MS) was used to probe the copper quota of fungal cells grown in defined medium and recovered from brain, which suggested a role for a copper-protective function of CTR4 in combination with cell metallothioneins under copper-replete conditions. In summary, these data suggest a role for CTR4 in copper-related homeostasis and subsequently in fungal virulence. IMPORTANCE:Crytococcus neoformans is a significant global fungal pathogen, and copper homeostasis is a relatively unexplored aspect of microbial pathogenesis that could lead to novel therapeutics. Previous studies correlated expression levels of a Ctr4 copper transporter to development of meningoencephalitis in a patient cohort of solid-organ transplants, but a direct role for Ctr4 in mammalian pathogenesis has not been demonstrated. The present studies utilize a ?ctr4 mutant strain which revealed an important role for CTR4 in C. neoformans infections in mice and relate the gene product to homeostatic control of copper and growth under nutrient-restricted conditions. Robust expression levels of CTR4 during fungal infection were exploited to demonstrate expression and lung cryptococcal disease using ex vivo fluorescence imaging. In summary, these studies are the first to directly demonstrate a role for a copper transporter in fungal disease and provide an ex vivo imaging tool for further study of cryptococcal gene expression and pathogenesis.

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Role of actin-bundling protein Sac6 in growth of Cryptococcus neoformans at low oxygen concentration.

Publications

Role of actin-bundling protein Sac6 in growth of Cryptococcus neoformans at low oxygen concentration.

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