Figure 3.

The dia2 ΔF-box mutant exhibits growth and S-phase defects. (A) The ΔF-box mutant is cold- and HU-sensitive. The dia2 strain was transformed with DIA2, ΔF-box and empty vector DNA. Serial dilutions of each were then spotted on rich media containing the indicated amount of HU and incubated at either 25 or 30°C. (B) The ΔF-box mutant enters S-phase earlier than WT cells. WT and dia2 ΔF-box cells were arrested in G1 with alpha factor and then released from arrest at 25°C. Samples were taken at the indicated time points and used for flow cytometry. The percentage of S-phase cells was calculated for the 35-, 40-, 45-, and 50-min time points using FlowJo software and is shown next to the profile for each strain. (C) The dia2 ΔF-box mutants exhibit early bud emergence. Cells were treated as in B and budding morphology was assayed by light microscopy. At least 100 cells were counted per time point and the experiment was repeated three times. Representative results from a single experiment are shown. The identity of each strain and bud stage is indicated next to the graph.