Cell cycle regulation of Cln3.

<p>(A) Cln3 levels peak in mitosis. Cells expressing Cln3-13Myc were synchronized in G1 with alpha-factor and released into the cell cycle. Samples were collected for flow cytometry and Western blot analysis at 15-minute intervals. Western blots for Cln3-13Myc, Clb2 and Cdc28 are shown. Cell cycle position was monitored by flow cytometry, right. Quantification of Cln3 protein levels is shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002851#pgen.1002851.s001" target="_blank">Figure S1A</a>. (B) Cln3 is unstable in G1 and mitosis. Cycloheximide-chase assay showing levels of Cln3-13Myc and Cdc28 in cells arrested in G1 with alpha-factor, or in mitosis (M) with nocodazole, and then treated with cycloheximide for the indicated number of minutes (min chx). Cell cycle profiles are shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002851#pgen.1002851.s001" target="_blank">Figure S1B</a>. (C) Cln3 stability is regulated by Cdc53. Cycloheximide-chase assay showing levels of Cln3-13Myc and Cdc28 in wild-type cells (<i>CDC53</i>) and <i>cdc53-1</i> temperature-sensitive cells before (23°C) and after (37°C) incubation at the non-permissive temperature. Following incubation at 37°C, cycloheximide was added for the indicated number of minutes. Cell cycle profiles are shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002851#pgen.1002851.s001" target="_blank">Figure S1C</a>. For all gels, molecular weight markers are indicated at the left.