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Rapid redistribution of phosphatidylinositol-(4,5)-bisphosphate and septins during the Candida albicans response to caspofungin.

ABSTRACT: We previously showed that phosphatidylinositol-(4,5)-bisphosphate [PI(4,5)P2] and septin regulation play major roles in maintaining Candida albicans cell wall integrity in response to caspofungin and other stressors. Here, we establish a link between PI(4,5)P2 signaling and septin localization and demonstrate that rapid redistribution of PI(4,5)P2 and septins is part of the natural response of C. albicans to caspofungin. First, we studied caspofungin-hypersusceptible C. albicans irs4 and inp51 mutants, which have elevated PI(4,5)P2 levels due to loss of PI(4,5)P2-specific 5'-phosphatase activity. PI(4,5)P2 accumulated in discrete patches, rather than uniformly, along surfaces of mutants in yeast and filamentous morphologies, as visualized with a green fluorescent protein (GFP)-pleckstrin homology domain. The patches also contained chitin (calcofluor white staining) and cell wall protein Rbt5 (Rbt5-GFP). By transmission electron microscopy, patches corresponded to plasma membrane invaginations that incorporated cell wall material. Fluorescently tagged septins Cdc10 and Sep7 colocalized to these sites, consistent with well-described PI(4,5)P2-septin physical interactions. Based on expression patterns of cell wall damage response genes, irs4 and inp51 mutants were firmly positioned within a group of caspofungin-hypersusceptible, septin-regulatory protein kinase mutants. irs4 and inp51 were linked most closely to the gin4 mutant by expression profiling, PI(4,5)P2-septin-chitin redistribution and other phenotypes. Finally, sublethal 5-min exposure of wild-type C. albicans to caspofungin resulted in redistribution of PI(4,5)P2 and septins in a manner similar to those of irs4, inp51, and gin4 mutants. Taken together, our data suggest that the C. albicans Irs4-Inp51 5'-phosphatase complex and Gin4 function upstream of PI(4,5)P2 and septins in a pathway that helps govern responses to caspofungin.

SUBMITTER: Badrane H

PROVIDER: S-EPMC3421880 | biostudies-other | 2012 Sep

REPOSITORIES: biostudies-other

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Rapid redistribution of phosphatidylinositol-(4,5)-bisphosphate and septins during the Candida albicans response to caspofungin.

Badrane Hassan H Nguyen M Hong MH Blankenship Jill R JR Cheng Shaoji S Hao Binghua B Mitchell Aaron P AP Clancy Cornelius J CJ

Antimicrobial agents and chemotherapy 20120611 9

We previously showed that phosphatidylinositol-(4,5)-bisphosphate [PI(4,5)P2] and septin regulation play major roles in maintaining Candida albicans cell wall integrity in response to caspofungin and other stressors. Here, we establish a link between PI(4,5)P2 signaling and septin localization and demonstrate that rapid redistribution of PI(4,5)P2 and septins is part of the natural response of C. albicans to caspofungin. First, we studied caspofungin-hypersusceptible C. albicans irs4 and inp51 m ...[more]

PMID: 22687514

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Project description:Phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] activates the yeast cell wall integrity pathway. Candida albicans exposure to caspofungin results in the rapid redistribution of PI(4,5)P2 and septins to plasma membrane foci and subsequent fungicidal effects. We studied C. albicans PI(4,5)P2 and septin dynamics and protein kinase C (PKC)-Mkc1 cell wall integrity pathway activation following exposure to caspofungin and other drugs. PI(4,5)P2 and septins were visualized by live imaging of C. albicans cells coexpressing green fluorescent protein (GFP)-pleckstrin homology (PH) domain and red fluorescent protein-Cdc10p, respectively. PI(4,5)P2 was also visualized in GFP-PH domain-expressing C. albicans mkc1 mutants. Mkc1p phosphorylation was measured as a marker of PKC-Mkc1 pathway activation. Fungicidal activity was assessed using 20-h time-kill assays. Caspofungin immediately induced PI(4,5)P2 and Cdc10p colocalization to aberrant foci, a process that was highly dynamic over 3 h. PI(4,5)P2 levels increased in a dose-response manner at caspofungin concentrations of ?4× MIC and progressively decreased at concentrations of ?8× MIC. Caspofungin exposure resulted in broad-based mother-daughter bud necks and arrested septum-like structures, in which PI(4,5)P2 and Cdc10 colocalized. PKC-Mkc1 pathway activation was maximal within 10 min, peaked in response to caspofungin at 4× MIC, and declined at higher concentrations. The caspofungin-induced PI(4,5)P2 redistribution remained apparent in mkc1 mutants. Caspofungin exerted dose-dependent killing and paradoxical effects at ?4× and ?8× MIC, respectively. Fluconazole, amphotericin B, calcofluor white, and H2O2 did not impact the PI(4,5)P2 or Cdc10p distribution like caspofungin did. Caspofungin exerts rapid PI(4,5)P2-septin and PKC-Mkc1 responses that correlate with the extent of C. albicans killing, and the responses are not induced by other antifungal agents. PI(4,5)P2-septin regulation is crucial in early caspofungin responses and PKC-Mkc1 activation.

Dataset Information

Rapid redistribution of phosphatidylinositol-(4,5)-bisphosphate and septins during the Candida albicans response to caspofungin.

Publications

Rapid redistribution of phosphatidylinositol-(4,5)-bisphosphate and septins during the Candida albicans response to caspofungin.

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