Figure 2.
Inducible Transcription Is Impaired in Oversized Cells
(A–D) Imaging of cells released from a cdc28-13 block expressing Whi5-tdTomato and (A, B) CLN2pr-GFP or (C, D) CLB2pr-GFP into medium containing 2% glucose lacking drugs. Mean GFP intensities were measured on maximal projections and corrected for background and autofluorescence. (A) Traces are aligned when nuclear export of Whi5 was completed. Asterisks indicate p < 0.05 (Mann-Whitney U test). (E–G) cdc28-4 cells were arrested at 35°C as indicated and transcription was induced by addition of galactose or alpha factor (αF). mRNA concentration was determined by (E, F) RT-qPCR relative to ACT1 mRNA or by (G) microarray analysis 0 min and 40 min after αF exposure. Genes induced more than 4-fold in wild-type cells were quantified (27 genes). Asterisks indicate p < 0.01 (Wilcoxon matched-pairs signed rank test). (H) Chromatin immuno-precipitation before and 30 min after galactose addition in arrested cdc28-13 cells, expressing either Gal4-3V5 or 3V5-Gal80. (I) Western blot of phosphorylated Fus3 (P-Fus3) and total Fus3 in cdc28-4 G1 arrested cells 15 min after pheromone exposure. Kar2 was used as a loading control. Asterisks mark P-Fus3 and Fus3. Note: Fus3 phosphorylation occurs most efficiently during G1. Fus3-P in asynchronously growing cells (lane 3) is therefore lower than in small G1 arrested cells (2 h arrest, lane 4).