Figure 6

(a) Growth of CTNS-Δgydql transformed in different single delete and double delete vps mutants on medium containing cystine. Different vacuolar protein sorting defective mutants transformed with, CTNS-Δgydql expressed under TEF promoter, and examined by dilution spotting on minimal media containing 200 µM Methionine, & different concentration of cystine as described in experimental procedures. The photographs were taken after 2–3 days of incubation at 30 °C. The experiment was repeated with three independent transformations. (b) Comparison of uptake in vps1Δ, vps1Δvps17Δ and vps1Δssh4Δ backgrounds. For quantification of functional activity of the CTNS constructs in different backgrounds, the initial rate of ³⁵S-cystine uptake was measured in S. cerevisiae vps1Δ, vps1Δvps17Δ and vps1Δssh4Δ transformed with CTNS-WT, CTNS-Δgydql in presence of 50 µM cystine. The cells were harvested at 10 and 20 min. intervals and results were plotted as nmol/mg protein/min after deducting the background uptake. The results represent the means of two different experiments, each in duplicate (±S.D., n = 4).