MCC/eisosome proteins Sur7, Pil1, and Lsp1 are important for resistance of <i>Candida albicans</i> to copper.

2019-01-11T18:38:04Z (GMT) by Lois M. Douglas James B. Konopka
<p>(A) <i>C</i>. <i>albicans</i> strains lacking the indicated MCC/eisosome genes were replica-plated onto minimal medium containing either 0 μM or 500 μM CuSO<sub>4</sub> and incubated at 30°C for 48 hr. Only the <i>sur7Δ</i> and the <i>pil1Δ lsp1Δ</i> double deletion demonstrated strong sensitivity. (B) <i>SUR7</i> overexpression partially rescued the copper sensitivity of the <i>pil1Δ lsp1Δ</i> strain. <i>pil1Δ lsp1Δ</i> strains containing zero, one, or two extra copies of the <i>SUR7</i> gene were spotted onto minimal medium in the absence or presence of 1 mM CuSO<sub>4</sub> and incubated for 48 hr at 30°C. (C) The highest concentration of copper at which the indicated strains could grow after incubation in synthetic medium in 96-well plates for 48 hr at 37°C with different concentrations of copper. The <i>sur7Δ</i>, <i>pil1Δ lsp1Δ</i>, and known copper sensitive mutant <i>cup1Δ crp1Δ</i> could only grow at lower concentrations of copper than the wild-type strain, indicating increased sensitivity. (D) Time course study of viability after exposure to copper. Cells were incubated in H<sub>2</sub>O or 0.5 μM CuSO<sub>4</sub> at 37°C for the indicated time and then an aliquot was spread on YPD medium to determine viable colony forming units (CFU). All deletion mutants exhibited lower viability after incubation in CuSO<sub>4</sub>. Images in panel A are representative of three independent experiments, and those in panel B of two independent experiments, all performed on different days. The results in panel C represent averages of three independent experiments and those in panel D of four independent experiments, all performed on different days. Error bars indicate SD. ****, P<0.001. ***, P<0.01. **, P<0.01 by two-way ANOVA. Strains used were: WT, wild-type DIC185; <i>sur7Δ</i> (YJA11); <i>sur7Δ</i> comp. (YJA12); <i>pil1Δ lsp1Δ</i> (YHXW21-1); <i>pil1Δ lsp1Δ</i> comp (+ <i>LSP1</i>, YHXW23-1); <i>seg1Δ</i> (YLD182-9-10-2); <i>slm2Δ</i> (YHXW36-1); <i>pkh1Δ pkh3Δ/PKH3</i> (YLD146-11-2-5); <i>pst1Δ pst2Δ pst3Δ ycp4Δ</i> (LLF060); <i>nce102Δ</i> (YHXW14); <i>pun1Δ</i> (YLD204-2); <i>fmp45Δ</i> (YHXW3), <i>pil1Δ lsp1Δ</i> + 1x<i>SUR7</i> (YHXW45-1), <i>pil1Δ lsp1Δ</i> + 2x<i>SUR7</i> (YHXW46-1), and <i>cup1Δ crp1Δ</i> (KC25).</p>