# Longer Ubiquinone Side Chains Contribute to Enhanced Farnesol Resistance in Yeasts
## 3 Results
### Farnesol Resistance in S. cerevisiae Depends on the UQ Isoprenologue, UQ6 vs. UQ9  
The importance of ubiquinone chain length for farnesol resistance was tested by comparing strains of _S. cerevisiae_ which produce UQ6 (wild type BY4741 and _\(\Delta\)coq1::_pYES + COQ1) and UQ9 (\(\Delta\)coq1::_pYES + _At2g34630_) [21]. Growth levels were observed in the presence and absence of 50-400 \(\upmu\)M farnesol (Figures 3 and 4). The two sets of experiments differ primarily in the degree of aeration achieved. Figure 3 describes high aeration growth, 50 mL of YPD in 250 mL Erlenmeyer flasks shaking continuously at 225 rpm, whereas Figure 4 describes low aeration growth, 0.2 mL of YPD per well in 96-well plates shaken for one min every 10 min. The 96-well plate format is technically advantageous for measuring the growth of triplicate samples at 10 min intervals over 25 h while using minimal amounts of farnesol. Additionally, the lower aeration shifts cellular metabolism towards a greater reliance on fermentation, allowing us to confirm our expectation that farnesol inhibits respiration [6,10] but not fermentation [27]. Our high aeration results in YPD (Figure 3) showed 3-4 fold increased resistance to farresol in the UQ9 producing strain compared with its UQ6 producing parent. This increased resistance to 50 and 100 \(\upmu\)M farresol for the UQ9-producing strain was evident in both the higher cell yields achieved (compare Figure 3b vs. Figure 3a) and the lower percentage of dead cells observed (compare Figure 3d vs. Figure 3c). Very similar results were obtained when the two yeast strains were grown in three defined, minimal growth media, namely glucose-salts-biotin (GSB), yeast nitrogen base (YNB), and glucose-phosphate-proline (GPP). In each case the BY4741 parent (UQ6) grew poorly in 50 \(\upmu\)M farresol and poorly (GSB) or not at all (YNB and GPP) in 100 \(\upmu\)M farresol whereas the UQ9-producing strain grew equally as well with and without 50 \(\upmu\)M farresol and 50-70% as well with 100 \(\upmu\)M farresol (data not shown). Similar results were observed for experiments carried out in 96-well plates under low aeration except that the yeast cells were less sensitive to farresol. In this case the UQ6 yeasts were inhibited only slightly by 50 \(\upmu\)M farresol (Figure 4B) and the cell yield achieved by UQ6 yeasts in 100 \(\upmu\)M farresol (Figure 4B) was 60% of the UQ9 yield (Figure 4C). These results are consistent with the low aeration conditions shifting cellular metabolism from respiration to fermentation. They reinforce the view that farresol itself is not toxic to yeast cells, but instead it is the ROS generated by farresol's interaction with respiring mitochondria which is toxic. Taken together these results agree with those of [PERSON] et al. [32] that under most growth conditions the length of the UQ tail makes very little difference but in the presence of farresol (Figures 3 and 4) it makes a substantial difference. Figure 2: Ubiquinone analysis in physiologically distinct _C. albicans_ morphological types. The analysis was undertaken by high-performance liquid chromatography (HPLC) in diode array detection (DAD) mode using UQ10 as the internal standard (IS) [21]. (**a**) Representative histogram from ubiquinone profile of _C. albicans_ cells grown in synthetic (glucose-salts-biotin, GSB) medium. (**b**) Quantification of ubiquinones as detected by HPLC-DAD. Trace indicates that the ubiquinone detected was below the sensitivity limit of 0.1 pmol unit\({}^{-1}\) OD600 nm. \({}^{*}\) Glucose-proline-phosphate (GPP) medium is supplemented with N-acetylglucosamine as an inducer of hyphal growth. GPD is the defined medium developed by [PERSON] et al. [27] for anaerobic growth of _C. albicans_. Data are means \(\pm\) standard deviation (SD) of duplicate measurements. Figure 4: _Cont_. Figure 3: Longer ubiquinones increase the farnesol survival in _S. cerevisiae_—high aeration. Growth curves in yeast extract-peptone-dextrose (YPD) of (**a**) _S. cerevisiae_ BY4741 and (**b**) _S.