17 research outputs found
Structure of Ycf1 in the dephosphorylated state
Structure of the Ycf1 transporter in the dephosphorylated state. </p
An Assessment of Growth Media Enrichment on Lipid Metabolome and the Concurrent Phenotypic Properties of <i>Candida albicans</i>
<div><p>A critical question among the researchers working on fungal lipid biology is whether the use of an enriched growth medium can affect the lipid composition of a cell and, therefore, contribute to the observed phenotypes. One presumption is that enriched medias, such as YPD (yeast extract, peptone and dextrose), are likely to contain lipids, which may homogenize with the yeast lipids and play a role in masking the actual differences in the observed phenotypes or lead to an altered phenotype altogether. To address this issue, we compared the lipids of <i>Candida albicans</i>, our fungus of interest, grown in YPD or in a defined media such as YNB (yeast nitrogen base). Mass spectrometry-based lipid analyses showed differences in the levels of phospholipids, including phosphatidylinositol, phosphatidylglycerol, lyso-phospholipids; sphingolipids, such as mannosyldiinositolphosphorylceramide; and sterols, such as ergostatetraenol. Significant differences were observed in 70 lipid species between the cells grown in the two media, but the two growth conditions did not affect the morphological characteristics of <i>C. albicans</i>. The lipid profiles of the YNB- and YPD-grown <i>C. albicans</i> cells did vary, but these differences did not influence their response to the majority of the tested agents. Rather, the observed differences could be attributed to the slow growth rate of the <i>Candida</i> cells in YNB compared to YPD. Notably, the altered lipid changes between the two media did impact the susceptibility to some drugs. This data provided evidence that changes in media can lead to certain lipid alterations, which may affect specific pathways but, in general, do not affect the majority of the phenotypic properties of <i>C. albicans</i>. It was determined that either YNB or YPD may be suitable for the growth and lipid analysis of <i>C. albicans</i>, depending upon the experimental requirements, but additional precautions are necessary when correlating the phenotypes with the lipids.</p></div
PGL composition of <i>C. albicans</i> cells grown in different medium.
<p>A) Total PGLs (as normalized total PGL+SL+SE mass spectral signal). B) Relative abundance of PGL classes (as % of normalized total PGL+SL+SE mass spectral signal). Values are mean of 2 independent analyses (n = 2). Data can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113664#pone.0113664.s001" target="_blank">Supplementary Sheet S1</a>. “*” indicates that <i>p</i>-value is <0.05.</p
Molecular lipid species composition of <i>C. albicans</i> cells grown in YPD or YNB.
<p>Data is represented as % of total PGL+SL+SE mass spectral signal normalized to the internal standards. Values are means ± SEM (n = 2). Data can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113664#pone.0113664.s001" target="_blank">Supplementary Sheet S1</a>. Only significant changes where <i>p</i>-value is <0.05 are depicted in this figure.</p
Growth phenotypes of <i>C. albicans</i> cells grown in YPD or YNB.
<p>A) Growth curve of CAI-4 cells grown in YPD and YNB medium. B) Generation time of CAI-4 cells grown in YPD or YNB medium. C) Cell morphology of CAI-4 cells grown in YPD or YNB medium. D) Spot assays showing growth of CAI-4 cells grown in YPD or YNB medium at variable temperatures. E) Hyphae formation of CAI-4 cells grown in YPD or YNB. <i>Candida</i> strains were cultured in YPD or YNB medium at 30°C as described in previously <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113664#pone.0113664-Singh1" target="_blank">[11]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113664#pone.0113664-Shah1" target="_blank">[18]</a>. To test the hyphae formation YPD or YNB grown cells were re-grown on spider agar plates at 37°C for 5 days. Values are means ± SEM (n = 3). “*” indicates that p-value is <0.05.</p
Effect of cell wall perturbing agents and protein biosynthesis inhibitors on <i>C. albicans</i> cells grown in YPD or YNB.
<p>A) CW. B) CR. C) TX-100. D) SDS. E) CYCLO. Values are means ± SEM (n = 4). YPD and YNB control datasets are same for the subfigures A–E.</p
PCA analysis of lipid species of <i>C. albicans</i> cells grown in YPD or YNB.
<p>The figure shows the 2D- PCA score plots, where the scores for the first three principal components, explaining >40% of the variance, are plotted. A) Principal component 1 versus Principal component 2. B) Principal component 1 versus Principal component 3. Each point in the PCA plot represents the principal component score of the individual replicate. Data for the PCA analyses was taken from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113664#pone.0113664.s001" target="_blank">Supplementary Sheet S1</a>. Loading values are indicated in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113664#pone-0113664-t001" target="_blank">Table 1</a>.</p
Effect of lipid biosynthesis inhibitors on <i>C. albicans</i> cells grown in YPD or YNB.
<p>A) Effect of PC analogue: MLT. B) Effect of phytoceramide biosynthesis inhibitor: MYR. C–D) Effect of ergosterol binding drugs: AMPB and NYS. E–F) Effect of ergosterol biosynthesis inhibitors: FLC, MICO, KETO and ITRA. Values are means ± SEM (n = 4). YPD and YNB control datasets are same for the subfigures A–H.</p
Loadings of principal components 1 and 2 from PCA analysis of lipids species of <i>C. albicans</i> cells grown in different medium.
<p>The 12 highest and 12 lowest values are indicated.</p><p>Loadings of principal components 1 and 2 from PCA analysis of lipids species of <i>C. albicans</i> cells grown in different medium.</p
Sterol composition of <i>C. albicans</i> cells grown in YPD or YNB.
<p>A) Total SEs (as normalized total PGL+SL+SE mass spectral signal). B) Relative abundance of SE classes (as % of normalized total PGL+SL+SE mass spectral signal). Values are mean of 2 independent analyses (n = 2). Data can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0113664#pone.0113664.s001" target="_blank">Supplementary Sheet S1</a>. “*” indicates that <i>p</i>-value is <0.05.</p