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. Author manuscript; available in PMC: 2015 Sep 11.
Published in final edited form as: Cell. 2014 Sep 11;158(6):1375–1388. doi: 10.1016/j.cell.2014.07.050

Figure 5. Cdc48p-dependent function of Otu1p in vivo.

Figure 5

(A) The degradation of a fusion of sCPY* with DHFR and a hemagglutinin (HA) tag (sCPY*-DHFR-HA) was tested in S. cerevisiae. The cells were transformed with an empty vector or plasmids expressing FLAG-tagged wild type or mutant Otu1p (Otu1p (C120S)) from a Gal1 promoter. Where indicated, Otu1p variants lacking their Ubx domains were expressed instead. The samples were analyzed at different time points after addition of cycloheximide (chx) by SDS-PAGE and immunoblotting with anti-HA and anti-FLAG antibodies. Loading controls were performed with Kar2p antibodies.

(B) Quantification of two experiments as in (A) (means and standard deviations).

(C) As in (A), but following simultaneously the degradation of Erg1p and Deg1-LacZ with antibodies to the endogenous protein and to LacZ, respectively.

(D) Quantification of three experiments as in (C) (means and standard deviations).

(E) The degradation of sCPY*-DHFR-HA was analyzed in cells lacking Otu1p and wild type (wt) cells. Cells lacking Hrd1p were analyzed in parallel.

(F) Quantification of the experiment in (E).

See also Figure S5.