Function of C. albicans Ess1 linker mutants in S. cerevisiae.

Host strain (MATa ura3-1 leu2-3, 112 trp1-1 can1-100 ade2-1 his3-11, 15 [phi+] ess1ΔTRP1+YEpESS1) is an ess1ΔTRP1 mutant of S. cerevisiae (Wu and Hanes, 2000, unpublished) covered by a 2 µM, URA3 plasmid expressing ESS1 (YEpEss1). Cells were transformed with the indicated plasmids (all 2 µM, HIS3) and plated on complete synthetic media (CSM) minus uracil (ura), histidine (his) and tryptophan (trp). Colonies were picked and passaged (20 ul into 3ml) for three successive overnights in liquid CSM minus his. After 3 days, individual colonies were patched onto CSM minus his trp plates, and replica-plated to CSM minus ura (for Ura+), 1mg/ml 5-FOA (for Ura−), and CSM minus his trp (for total patch number) and scored for uracil prototrophy after 1 day. For the helix subsitution mutants, three independent clone isolates of helix substitution plasmid, pDS413(pm) were tested. For the linker-swapped mutants, two independent clone isolates of plasmid, pDS413(sw) were tested.