Fig. 2. Mek1 is a meiosis-specific nuclear protein. (A) Schematic representation of pat1-driven synchronous meiosis. Vegetatively growing cells are blocked in G1 by nitrogen starvation during 14 hours and then induced to enter meiosis synchronously by inactivating the Pat1 kinase at 34°C (time 0 in B, C and D). The approximate timing of the major meiotic landmarks, as determined in (B), is indicated. (B) Synchronous meiosis of strain S964. Left panel, DNA content measured by FACS analysis. The period in which premeiotic DNA replication takes place is indicated (meiS). Note that the 1C peak that appears after 7 hours corresponds to free spores that are released from asci owing to sonication during the preparation of cells for FACS. Right panel, meiotic progression was followed by DAPI staining of nuclei and sporulation by microscopic observation of asci. The peaks of meiosis I (MI), meiosis II (MII) and spore formation (Spo) are indicated. (C) Northern blot analysis of mek1⁺ expression during the synchronous meiosis of strain S964 shown in B. 18S rRNA levels are shown as a loading control. (D) Western blot analysis of Mek1-HA production during a synchronous meiosis in strain S1294. Tubulin is presented as a loading control. (E) Immunofluorescence analysis of cells from strain S1294 (mek1-HA), after 3 hours of induction of meiosis, stained with DAPI (blue) and anti-HA antibodies (red). The merged image is presented in the right column. Three representative cells are shown.