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HapX positively and negatively regulates the transcriptional response to iron deprivation in Cryptococcus neoformans.

Jung WH, Saikia S, Hu G, Wang J, Fung CK, D'Souza C, White R, Kronstad JW - PLoS Pathog. (2010)

Bottom Line: However, we also found that HapX plays both positive and negative roles in the regulation of gene expression, including a positive regulatory role in siderophore transporter expression.Although both hapX and hap3 mutants were defective in heme utilization in culture, only HapX made a contribution to virulence, and loss of HapX in a strain lacking the high-affinity iron uptake system did not cause further attenuation of disease.Overall, these results indicated that C. neoformans employs multiple strategies for iron acquisition during infection.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology, Chung-Ang University, Gyeonggi-Do, Republic of Korea.

ABSTRACT
The fungal pathogen Cryptococcus neoformans is a major cause of illness in immunocompromised individuals such as AIDS patients. The ability of the fungus to acquire nutrients during proliferation in host tissue and the ability to elaborate a polysaccharide capsule are critical determinants of disease outcome. We previously showed that the GATA factor, Cir1, is a major regulator both of the iron uptake functions needed for growth in host tissue and the key virulence factors such as capsule, melanin and growth at 37°C. We are interested in further defining the mechanisms of iron acquisition from inorganic and host-derived iron sources with the goal of understanding the nutritional adaptation of C. neoformans to the host environment. In this study, we investigated the roles of the HAP3 and HAPX genes in iron utilization and virulence. As in other fungi, the C. neoformans Hap proteins negatively influence the expression of genes encoding respiratory and TCA cycle functions under low-iron conditions. However, we also found that HapX plays both positive and negative roles in the regulation of gene expression, including a positive regulatory role in siderophore transporter expression. In addition, HapX also positively regulated the expression of the CIR1 transcript. This situation is in contrast to the negative regulation by HapX of genes encoding GATA iron regulatory factors in Aspergillus nidulans and Schizosaccharomyces pombe. Although both hapX and hap3 mutants were defective in heme utilization in culture, only HapX made a contribution to virulence, and loss of HapX in a strain lacking the high-affinity iron uptake system did not cause further attenuation of disease. Therefore, HapX appears to have a minimal role during infection of mammalian hosts and instead may be an important regulator of environmental iron uptake functions. Overall, these results indicated that C. neoformans employs multiple strategies for iron acquisition during infection.

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Related in: MedlinePlus

Acidic pH rescues the growth of double mutants on ferric chloride and hemin.Ten-fold serial dilutions of cells (starting at 104 cells) were spotted onto solid YPD medium, low-iron medium (YNB+100 µM BPS) at either pH 5.0 or 7.0, and low-iron medium supplemented with 100 µM FeCl3, 10 µM Hemin or 10 µM Feroxamine at each of the two pH conditions. The plates were incubated at 30°C for two days.
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ppat-1001209-g009: Acidic pH rescues the growth of double mutants on ferric chloride and hemin.Ten-fold serial dilutions of cells (starting at 104 cells) were spotted onto solid YPD medium, low-iron medium (YNB+100 µM BPS) at either pH 5.0 or 7.0, and low-iron medium supplemented with 100 µM FeCl3, 10 µM Hemin or 10 µM Feroxamine at each of the two pH conditions. The plates were incubated at 30°C for two days.

Mentions: Possibilities for additional iron uptake mechanisms include low-affinity or ferrous iron-specific systems that may respond to iron mobilized by reduced pH during infection. A reduction in pH increases iron availability and acidic metabolites such as acetate are known to accumulate during cryptococcal infection [36]. In addition, a low-affinity uptake system for inorganic iron was previously identified in C. neoformans by physiological experiments, and intracellular residence of fungal cells in the acidic phagolysosomal compartment may also influence iron availability [37]. We tested the possibility of an additional uptake function by reexamining the growth of the hap mutants alone or in combination with the cfo1Δ mutation on medium at reduced pH (Figure 9). The plate assays revealed that a reduction in pH to 5.0 rescued the growth of the double mutants on both ferric chloride and hemin in support of the idea that iron availability is enhanced under acidic conditions to allow uptake via an alternative mechanism(s). The results also suggest that the cells require Hap functions to utilize both iron sources in the situation where the loss of Cfo1 activity causes a defect in the high-affinity, reductive iron uptake pathway.


HapX positively and negatively regulates the transcriptional response to iron deprivation in Cryptococcus neoformans.

Jung WH, Saikia S, Hu G, Wang J, Fung CK, D'Souza C, White R, Kronstad JW - PLoS Pathog. (2010)

Acidic pH rescues the growth of double mutants on ferric chloride and hemin.Ten-fold serial dilutions of cells (starting at 104 cells) were spotted onto solid YPD medium, low-iron medium (YNB+100 µM BPS) at either pH 5.0 or 7.0, and low-iron medium supplemented with 100 µM FeCl3, 10 µM Hemin or 10 µM Feroxamine at each of the two pH conditions. The plates were incubated at 30°C for two days.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2991262&req=5

ppat-1001209-g009: Acidic pH rescues the growth of double mutants on ferric chloride and hemin.Ten-fold serial dilutions of cells (starting at 104 cells) were spotted onto solid YPD medium, low-iron medium (YNB+100 µM BPS) at either pH 5.0 or 7.0, and low-iron medium supplemented with 100 µM FeCl3, 10 µM Hemin or 10 µM Feroxamine at each of the two pH conditions. The plates were incubated at 30°C for two days.
Mentions: Possibilities for additional iron uptake mechanisms include low-affinity or ferrous iron-specific systems that may respond to iron mobilized by reduced pH during infection. A reduction in pH increases iron availability and acidic metabolites such as acetate are known to accumulate during cryptococcal infection [36]. In addition, a low-affinity uptake system for inorganic iron was previously identified in C. neoformans by physiological experiments, and intracellular residence of fungal cells in the acidic phagolysosomal compartment may also influence iron availability [37]. We tested the possibility of an additional uptake function by reexamining the growth of the hap mutants alone or in combination with the cfo1Δ mutation on medium at reduced pH (Figure 9). The plate assays revealed that a reduction in pH to 5.0 rescued the growth of the double mutants on both ferric chloride and hemin in support of the idea that iron availability is enhanced under acidic conditions to allow uptake via an alternative mechanism(s). The results also suggest that the cells require Hap functions to utilize both iron sources in the situation where the loss of Cfo1 activity causes a defect in the high-affinity, reductive iron uptake pathway.

Bottom Line: However, we also found that HapX plays both positive and negative roles in the regulation of gene expression, including a positive regulatory role in siderophore transporter expression.Although both hapX and hap3 mutants were defective in heme utilization in culture, only HapX made a contribution to virulence, and loss of HapX in a strain lacking the high-affinity iron uptake system did not cause further attenuation of disease.Overall, these results indicated that C. neoformans employs multiple strategies for iron acquisition during infection.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology, Chung-Ang University, Gyeonggi-Do, Republic of Korea.

ABSTRACT
The fungal pathogen Cryptococcus neoformans is a major cause of illness in immunocompromised individuals such as AIDS patients. The ability of the fungus to acquire nutrients during proliferation in host tissue and the ability to elaborate a polysaccharide capsule are critical determinants of disease outcome. We previously showed that the GATA factor, Cir1, is a major regulator both of the iron uptake functions needed for growth in host tissue and the key virulence factors such as capsule, melanin and growth at 37°C. We are interested in further defining the mechanisms of iron acquisition from inorganic and host-derived iron sources with the goal of understanding the nutritional adaptation of C. neoformans to the host environment. In this study, we investigated the roles of the HAP3 and HAPX genes in iron utilization and virulence. As in other fungi, the C. neoformans Hap proteins negatively influence the expression of genes encoding respiratory and TCA cycle functions under low-iron conditions. However, we also found that HapX plays both positive and negative roles in the regulation of gene expression, including a positive regulatory role in siderophore transporter expression. In addition, HapX also positively regulated the expression of the CIR1 transcript. This situation is in contrast to the negative regulation by HapX of genes encoding GATA iron regulatory factors in Aspergillus nidulans and Schizosaccharomyces pombe. Although both hapX and hap3 mutants were defective in heme utilization in culture, only HapX made a contribution to virulence, and loss of HapX in a strain lacking the high-affinity iron uptake system did not cause further attenuation of disease. Therefore, HapX appears to have a minimal role during infection of mammalian hosts and instead may be an important regulator of environmental iron uptake functions. Overall, these results indicated that C. neoformans employs multiple strategies for iron acquisition during infection.

Show MeSH
Related in: MedlinePlus