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. Author manuscript; available in PMC: 2011 May 12.
Published in final edited form as: Cell. 2010 Nov 12;143(4):579–591. doi: 10.1016/j.cell.2010.10.028

Figure 6. Mutations of ERAD components on the cytosolic side affect substrate-Hrd1p crosslinking.

Figure 6

(A) sCPY*-DHFR-HA with photoreactive probes at the indicated positions was expressed in cells together with Myc-tagged Hrd1 or a mutant defective in its ubiquitin ligase activity (C399S). Following UV irradiation, detergent-solubilized membranes were subjected to immunoprecipitation with HA antibodies, and bound proteins were analyzed by SDS-PAGE and immunoblotting with Myc antibodies. The numbers give percentage of crosslinking intensity relative to wild type and are the average of four experiments.

(B) As in (A), with either wild type (wt) or usa1Δ cells. The numbers are the average of two experiments.

(C) As in (A), with either wild type (wt) cells or cells bearing the indicated temperature-sensitive alleles of components of the Cdc48 ATPase complex analyzed after 1hr incubation at the restrictive temperature of 37°C. The numbers are the average of two experiments.

See also Figure S7.