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. 2008 Jun;19(6):2457–2464. doi: 10.1091/mbc.E08-02-0227

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© 2008 by The American Society for Cell Biology

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Figure 4. — Mdm30p, Cdc34p, and Cdc53p are required for K48-linked ubiquitylation of Fzo1p. (A) Cellular levels of Fzo1p, Mdm30p (Mdm30-HA), or a version of Mdm30p with a mutated F-box incapable of binding to Skp1p (fbox-HA) were analyzed in whole cell extracts prepared from the indicated strains (W303 background) grown at 30°C. Unmodified and ubiquitylated forms of Fzo1p are indicated by thick and thin arrows, respectively. PGK is shown as a loading control. (B) Ubiquitylated forms of Fzo1p were analyzed in whole cell extracts prepared from wild-type, cdc53ts, or cdc34ts cells grown at 23°C and shifted to 37°C. PGK serves as a loading control. (C) Anti-Fzo1p immunoblot of whole cell extracts prepared from fzo1Δ and wild-type cells (BY4742) transformed with vectors overexpressing either ubiquitin (WT), K48R ubiquitin, K63R ubiquitin, or empty vector as control.