Transcription profiling by array of Saccharomyces cerevisiae to study Met4 hyperactivation over a timecourse in met31 single-deletion and met32 single-deletion strains compared to Met31/Met32 wild-type strains
ABSTRACT: The met31 single delete and the met32 single delete strains were analyzed for genome-wide transcription at different times following Met4 hyeractivation and compared to cells that contain both Met31 and Met32.
Yeast sulfur metabolism is transcriptionally regulated by the activator Met4. Met4 lacks DNA-binding ability and relies on interactions with Met31 and Met32, paralogous proteins that bind the same cis-regulatory element, to activate its targets. Although Met31 and Met32 are redundant for growth in the absence of methionine, studies indicate that Met32 has a prominent role over Met31 when Met30, a negative regulator of Met4 and Met32, is inactive. To characterize different roles of Met31 and Met3 ...[more]
Project description:All yeast strains used in this study (Table 1) are in the W303 background (ade2-1 can1-100, his3-1,15 leu2-3,112 trp1-1 ura3). For sulfur limitation microarray studies, WT, met4 delete, met31 delete met32 delete, cbf1 delete, and met28 delete strains were grown in minimal B-media [see Cherest, H., and Surdin-Kerjan, Y. (1992). Genetic analysis of a new mutation conferring cysteine auxotrophy in Saccharomyces cerevisiae: updating of the sulfur metabolism pathway. Genetics 130, p51-58 for B-media composition] supplemented with 0.5mM methionine as the sole sulfur source. An aliquot of cells was harvested for a t=0 time point while the remainder were filtered through a .22um Stericup filter (Millipore), then washed and resuspended in pre-warmed (30 C) B-media lacking any source of sulfur. Cells were harvested after 20, 40, and 80 minutes.