Figure 2.

Genetic interaction between doa4Δ and doa3. (A) Enhanced growth defect of a doa3-1 doa4Δ double mutant. Doubly mutant doa3-1 doa4Δ cells, as well as the corresponding single mutants, were grown at 30°C and 35°C for one week. The wild-type parents grow equally well at both temperatures. (B) Suppression of the low molecular mass ubiquitin conjugates found in doa4Δ cells by impairment of the proteasome. Extracts from exponentially growing wild-type, doa3-1, doa4Δ, and doa3-1 doa4Δ cells were separated on an 18% polyacrylamide gel, blotted to a polyvinylidene difluoride membrane, and probed with an affinity-purified antiubiquitin antibody (a gift of Cecile Pickart, Department of Biochemistry, School of Public Health, Johns Hopkins University, Baltimore, MD). The positions of ubiquitin and the mono-ubiquitinated and di-ubiquitinated conjugates specific to doa4 cells are indicated.