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. 2010 Sep 15;21(18):3205–3219. doi: 10.1091/mbc.E09-06-0452

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© 2010 by The American Society for Cell Biology

This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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Figure 5. — Cdc17 is degraded primarily in the nucleus. (A) ABy150 (S288C), ABy152 (ubc4Δ), ABy574 (UBC4-NES-3HA), ABy577 (ubc5Δ, UBC4-NES-3HA), and GAP510 (ubc4Δ, ubc5Δ) were heat shocked as described in the Materials and Methods, aliquots were grown on YPD plates for 3 d at 30°C, and colonies were counted to determine the percentage of cells that were viable following heat shock. (B) ABy605 (mcm10-1, UBC4-GFP) and ABy660 (mcm10-1, UBC4-NES-GFP) were examined using fluorescence microscopy. Merge represents GFP and DAPI. Yellow dots represent colocalization of Ubc4-GFP and DAPI-stained nuclei. Red dots represent DAPI-stained nuclei in which Ubc4-GFP is not present. Black circles represent vacuoles. (C) Asynchronous cultures of ABy013 (mcm10-1), ABy342 (mcm10-1, ubc4Δ), ABy455 (mcm10-1, UBC4-NES-3HA), ABy363 (mcm10-1, ubc5Δ), and ABy359 (mcm10-1, rad6Δ) grown at 25°C were shifted to 37°C for 180 min. Cdc17–3HA and α-tubulin were analyzed by Western blot. (D) The graph shows Cdc17/tubulin ratios at each time point for each strain relative to mcm10-1 at time 0 (average of 3 separate experiments, bars represent mean ± SD).