Supplementary data for Ramsey et al.
Supplementary data for Ramsey et al.
Genetic analysis does not support a role for cullins in NEF4 function in DNA repair.
Figure S 1. A cdc53 strain is not UV sensitive. UV sensitivity of wild-type (CDC53), cdc53-1, rad23∆, and cdc53-1rad23∆ strains. CDC53 encodes the essential yeast cullin Cul1, a subunit in multiple E3 ubiquitin ligase complexes. This graph shows the relative number of colonies that grew on agar plates at 30° C following irradiation with the indicated doses of UV light. For each strain, the number of colonies on the unirradiated control was normalized to 100% survival. The temperature-sensitive cdc53-1 strain displayed no sensitivity to UV light when grown at either 30° C or the semi-permissive temperature of 33° C (not shown). In contrast to the effects seen with deletion of NEF4 subunit genes, a cdc53-1rad23∆ strain was significantly less UV sensitive than the rad23∆ strain. The basis for this suppression is not known, but could be due to improved cell survival resulting from cdc53-1-imposed cell cycle delay. This analysis demonstrates that CDC53 does not have a unique role in NER. As there is no enhanced UV sensitivity in the cdc53-1rad23∆ strain compared to the single mutants, we conclude that CDC53 does not function in the NEF4 pathway.
Figure S 2. rtt101∆ and ygr003w∆ strains are not UV sensitive. UV sensitivity of wild-type (WT), rtt101∆, ygr003w∆, rad23∆, ygr003w∆ rad23∆, and rtt101∆ rad23∆ strains was determined as described in the legend to Figure S1. The error bars represent the standard deviations determined from measurements made in triplicate. RTT101 (also known as CUL8) and YGR003W (also known as CUL3) encode the two nonessential yeast cullins. rtt101∆ and ygr003w∆ strains had wild-type sensitivity to UV light and did not affect the UV sensitivity caused by rad23∆. These cullins therefore do not appear to have a unique or essential role in UV repair.