Fig. 1: Cab1 mutants display growth defects and increased susceptibility toward ergosterol targeting drugs.

a Schematic diagram of Coenzyme A synthetic pathway and biological roles of the Acetyl-CoA. Pantothenate kinase (PanK) catalyzes the phosphorylation of pantothenic acid to form 4’-phosphopantothenic acid, the first step in the biosynthesis of CoA. Phosphopantothenoylcysteine synthase (PPCS) converts 4’-phosphopantothenic acid to phosphopantothenoylcysteine. Phosphopantothenoylcysteine decarboxylase (PPCD) catalyzes the decarboxylation of phosphopantothenoylcysteine to form pantetheine. Pantetheine kinase (PanK) catalyzes the phosphorylation of pantetheine to form dephospho-CoA. Dephospho-CoA kinase (DPCK) catalyzes the transfer of a phosphate group from ATP to dephospho-CoA, resulting in the formation of CoA. b- c The cab1 mutants display susceptibility toward drugs targeting ergosterol pathway and non-ergosterol targeting pathway. Yeast liquid growth curve assay with known AFDs targeting the ergosterol biosynthesis pathway was done using yeast cells harboring different CAB1 mutants. Cells were inoculated into 100 µL of YPD liquid media containing the antifungals in serial dilutions at 10 cells per µL ratio and incubated at 30 °C while cell growth was monitored by OD₆₀₀. % of cell growth of individual mutant strain in presence of AFDs was obtained compared to the cell growth in the vehicle control. Liquid growth assays were conducted in 3 replicates (n = 3) and the plotted graphs represent the average of 3 data sets. d Normalized relative MIC₅₀ of yeast strains expressing Cab1 variants against AFDs. MIC₅₀ values were determined on MIC₅₀ data obtained from the liquid growth assays presented in Fig. 1b, c. These values were calculated by dividing the MIC₅₀ value of the cab1 mutant or the wild type strain for the specified AFD by the MIC₅₀ value of the wild type strain. The plotted graph represents average of four data sets ± SD. e Complementation of the wild type CAB1 gene restores AFD resistance in the cab1 mutants to the similar level as that observed in wild-type cells. The cab1∆/w303-1B strain harboring 1) wild type CAB1, 2) cab1^mutant, or 3) cab1^mutant + wild type CAB1, were inoculated into synthetic glucose medium and grown overnight. Cells were harvested and re-suspended in 0.9% NaCl. Ten-fold serial dilutions of cells were spotted onto the YPD plates containing fluconazole (5 µg/mL), amphotericin B (0.25 µg/mL), and terbinafine (10 µg/mL), caspofungin (30 ng/mL), hygromycin B (20 µg/mL), or cycloheximide (100 ng/mL), and incubated at 30 °C for 3 days. The representative images are from two independent experiments, each performed in duplicates.