FIGURE 3.
Maf1-id S388A regulates pol III transcription normally during carbon source switching. A, recovery of Pol III transcription after growth in glycerol medium in the Maf1-id S388A strain. Yeast cultures were grown to early log phase (OD ∼0.5) in glucose-containing minimal media before carbon source switching and RNA extraction. Cells were harvested after exponential growth in glucose media (Glu exp) and after a 3 h switch into glycerol-containing media (Gly 3h). An additional aliquot of glycerol-switched cells was refed glucose-containing media and grown for a further 3 h (Glu 3h). For carbon source switching cells were harvested by centrifugation and washed once in warm media containing the new carbon source before further growth. RNA extraction and Northern analysis was done as in Fig. 1B. B, quantitation of Northern data from multiple independent experiments as described in Fig. 3A. Relative transcription (normalized pre-tRNALeu band intensities) was referenced to values determined for WT Maf1 grown in glucose-containing medium. C, phosphorylation of Maf1-id S388A after recovery from growth in glycerol. Strains were grown in glucose media, switched into glycerol media for 3 h and then switched back into glucose media as in Fig. 3A. Maf1 phosphorylated forms were resolved by Phos-tag SDS-PAGE as described in Fig. 1E.