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. 2000 May;11(5):1697–1708. doi: 10.1091/mbc.11.5.1697

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Copyright © 2000, The American Society for Cell Biology

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UP* degradation was independent of HRD1 and HRD3 but dependent on UBC7. (A) Cycloheximide–chase assay of strains expressing UP* in a wild-type (RHY1951), hrd1Δ (RHY2094), hrd3Δ (RHY1904), and ubc7Δ (RHY1900) genetic background. After addition of cycloheximide, lysates were prepared at the indicated times and immunoblotted with antiserum generated against the last 10 residues of uracil permease. (B) Levels of UP* ubiquitination correlated with its HRD-independent degradation. Cultures of strains expressing UP* in a wild-type (RHY1216), hrd1Δ (RHY1222), hrd2-1 (RHY1218), hrd3Δ (RHY1223), or ubc7Δ (RHY1221) genetic background were lysed, and UP* was immunoprecipitated with antibodies generated against the N-terminus of uracil permease. Ubiquitination of UP* was assayed by immunoblotting with an anti-ubiquitin antibody.