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Identification of Aph1, a Phosphate-Regulated, Secreted, and Vacuolar Acid Phosphatase in Cryptococcus neoformans
Cryptococcus neoformans strains isolated from patients with AIDS secrete acid phosphatase, but the identity and role of the enzyme(s) responsible have not been elucidated. By combining a one-dimensional electrophoresis step with mass spectrometry, a canonically secreted acid phosphatase, CNAG_02944...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Society of Microbiology
2014
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4172073/ https://www.ncbi.nlm.nih.gov/pubmed/25227465 http://dx.doi.org/10.1128/mBio.01649-14 |
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author | Lev, Sophie Crossett, Ben Cha, So Young Desmarini, Desmarini Li, Cecilia Chayakulkeeree, Methee Wilson, Christabel F. Williamson, P. R. Sorrell, Tania C. Djordjevic, Julianne T. |
author_facet | Lev, Sophie Crossett, Ben Cha, So Young Desmarini, Desmarini Li, Cecilia Chayakulkeeree, Methee Wilson, Christabel F. Williamson, P. R. Sorrell, Tania C. Djordjevic, Julianne T. |
author_sort | Lev, Sophie |
collection | PubMed |
description | Cryptococcus neoformans strains isolated from patients with AIDS secrete acid phosphatase, but the identity and role of the enzyme(s) responsible have not been elucidated. By combining a one-dimensional electrophoresis step with mass spectrometry, a canonically secreted acid phosphatase, CNAG_02944 (Aph1), was identified in the secretome of the highly virulent serotype A strain H99. We created an APH1 deletion mutant (Δaph1) and showed that Δaph1-infected Galleria mellonella and mice survived longer than those infected with the wild type (WT), demonstrating that Aph1 contributes to cryptococcal virulence. Phosphate starvation induced APH1 expression and secretion of catalytically active acid phosphatase in the WT, but not in the Δaph1 mutant, indicating that Aph1 is the major extracellular acid phosphatase in C. neoformans and that it is phosphate repressible. DsRed-tagged Aph1 was transported to the fungal cell periphery and vacuoles via endosome-like structures and was enriched in bud necks. A similar pattern of Aph1 localization was observed in cryptococci cocultured with THP-1 monocytes, suggesting that Aph1 is produced during host infection. In contrast to Aph1, but consistent with our previous biochemical data, green fluorescent protein (GFP)-tagged phospholipase B1 (Plb1) was predominantly localized at the cell periphery, with no evidence of endosome-mediated export. Despite use of different intracellular transport routes by Plb1 and Aph1, secretion of both proteins was compromised in a Δsec14-1 mutant. Secretions from the WT, but not from Δaph1, hydrolyzed a range of physiological substrates, including phosphotyrosine, glucose-1-phosphate, β-glycerol phosphate, AMP, and mannose-6-phosphate, suggesting that the role of Aph1 is to recycle phosphate from macromolecules in cryptococcal vacuoles and to scavenge phosphate from the extracellular environment. |
format | Online Article Text |
id | pubmed-4172073 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | American Society of Microbiology |
record_format | MEDLINE/PubMed |
spelling | pubmed-41720732014-10-06 Identification of Aph1, a Phosphate-Regulated, Secreted, and Vacuolar Acid Phosphatase in Cryptococcus neoformans Lev, Sophie Crossett, Ben Cha, So Young Desmarini, Desmarini Li, Cecilia Chayakulkeeree, Methee Wilson, Christabel F. Williamson, P. R. Sorrell, Tania C. Djordjevic, Julianne T. mBio Research Article Cryptococcus neoformans strains isolated from patients with AIDS secrete acid phosphatase, but the identity and role of the enzyme(s) responsible have not been elucidated. By combining a one-dimensional electrophoresis step with mass spectrometry, a canonically secreted acid phosphatase, CNAG_02944 (Aph1), was identified in the secretome of the highly virulent serotype A strain H99. We created an APH1 deletion mutant (Δaph1) and showed that Δaph1-infected Galleria mellonella and mice survived longer than those infected with the wild type (WT), demonstrating that Aph1 contributes to cryptococcal virulence. Phosphate starvation induced APH1 expression and secretion of catalytically active acid phosphatase in the WT, but not in the Δaph1 mutant, indicating that Aph1 is the major extracellular acid phosphatase in C. neoformans and that it is phosphate repressible. DsRed-tagged Aph1 was transported to the fungal cell periphery and vacuoles via endosome-like structures and was enriched in bud necks. A similar pattern of Aph1 localization was observed in cryptococci cocultured with THP-1 monocytes, suggesting that Aph1 is produced during host infection. In contrast to Aph1, but consistent with our previous biochemical data, green fluorescent protein (GFP)-tagged phospholipase B1 (Plb1) was predominantly localized at the cell periphery, with no evidence of endosome-mediated export. Despite use of different intracellular transport routes by Plb1 and Aph1, secretion of both proteins was compromised in a Δsec14-1 mutant. Secretions from the WT, but not from Δaph1, hydrolyzed a range of physiological substrates, including phosphotyrosine, glucose-1-phosphate, β-glycerol phosphate, AMP, and mannose-6-phosphate, suggesting that the role of Aph1 is to recycle phosphate from macromolecules in cryptococcal vacuoles and to scavenge phosphate from the extracellular environment. American Society of Microbiology 2014-09-16 /pmc/articles/PMC4172073/ /pubmed/25227465 http://dx.doi.org/10.1128/mBio.01649-14 Text en Copyright © 2014 Lev et al. http://creativecommons.org/licenses/by-nc-sa/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution-Noncommercial-ShareAlike 3.0 Unported license (http://creativecommons.org/licenses/by-nc-sa/3.0/) , which permits unrestricted noncommercial use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Lev, Sophie Crossett, Ben Cha, So Young Desmarini, Desmarini Li, Cecilia Chayakulkeeree, Methee Wilson, Christabel F. Williamson, P. R. Sorrell, Tania C. Djordjevic, Julianne T. Identification of Aph1, a Phosphate-Regulated, Secreted, and Vacuolar Acid Phosphatase in Cryptococcus neoformans |
title | Identification of Aph1, a Phosphate-Regulated, Secreted, and Vacuolar Acid Phosphatase in Cryptococcus neoformans |
title_full | Identification of Aph1, a Phosphate-Regulated, Secreted, and Vacuolar Acid Phosphatase in Cryptococcus neoformans |
title_fullStr | Identification of Aph1, a Phosphate-Regulated, Secreted, and Vacuolar Acid Phosphatase in Cryptococcus neoformans |
title_full_unstemmed | Identification of Aph1, a Phosphate-Regulated, Secreted, and Vacuolar Acid Phosphatase in Cryptococcus neoformans |
title_short | Identification of Aph1, a Phosphate-Regulated, Secreted, and Vacuolar Acid Phosphatase in Cryptococcus neoformans |
title_sort | identification of aph1, a phosphate-regulated, secreted, and vacuolar acid phosphatase in cryptococcus neoformans |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4172073/ https://www.ncbi.nlm.nih.gov/pubmed/25227465 http://dx.doi.org/10.1128/mBio.01649-14 |
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