The Identification of Downstream Targets of the Cell Integrity Pathway in Saccharomyces cerevisiae by Oligonucleotide Microarray Analysis
Hooker, Gillian W.
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The Saccharomyces cerevisiae Cell Integrity (Cl) pathway is a Mitogen Activated Protein (MAP) kinase pathway involved in cellular response to changes in environment and in cell cycle regulation. Responses generated by the CI pathway are not completely understood. Genome-wide gene expression in mutants of two genes in the pathway, the MAP kinase SIt2 and the transcription factor RIm 1, activated by Slt2, was examined to identify additional genes controlled by the pathway. Gene expression in mutants was compared to that in wild-type strains at 24º and after a 45 minute, 37º heat shock. Labeled mRNA was hybridized to Affymetrix S.cerevisiae oligonucleotide microarrays. Clustering was used to identify groups of genes regulated together, and additional computational tools were used to find known and unknown upstream regulatory regions. Expression of 116 genes was affected by the slt2 mutation. Of these genes, 37 had computer-identified regulatory regions for SBF, a second Slt2-regulated transcription factor, and 15 had Rlml regulatory regions. Expression of 57 genes was affected by the rim 1 mutation, 11 of which had SBF regions and 13 of which had RIm 1 regions. Thus, genes with predicted binding regions are indeed regulated by the pathway. That genes affected by these mutations lacked both SBF and Rlm 1 sites indicated the possible existence of an additional transcription factor. Searches for recurring sequence motifs in these genes identified new potential regulatory regions. Many genes in all regulated sets are involved in cell wall integrity, as expected for targets of the CI pathway. Investigation of non-cell wall genes may elucidate other aspects of the response and provide clues about similar response mechanisms in higher eukaryotic organisms.With honors.