We describe a genome-wide characterization of mRNA transcript levels in yeast grown on the fatty acid oleate, determined using Serial Analysis of Gene Expression (SAGE). Comparison of this SAGE library with that reported for glucose grown cells revealed the dramatic adaptive response of yeast to a change in carbon source. A major fraction (Ͼ20%) of the 15,000 mRNA molecules in a yeast cell comprised differentially expressed transcripts, which were derived from only 2% of the total number of ϳ6300 yeast genes. Most of the mRNAs that were differentially expressed code for enzymes or for other proteins participating in metabolism (e.g., metabolite transporters). In oleate-grown cells, this was exemplified by the huge increase of mRNAs encoding the peroxisomal ␤-oxidation enzymes required for degradation of fatty acids. The data provide evidence for the existence of redox shuttles across organellar membranes that involve peroxisomal, cytoplasmic, and mitochondrial enzymes. We also analyzed the mRNA profile of a mutant strain with deletions of the PIP2 and OAF1 genes, encoding transcription factors required for induction of genes encoding peroxisomal proteins. Induction of genes under the immediate control of these factors was abolished; other genes were up-regulated, indicating an adaptive response to the changed metabolism imposed by the genetic impairment. We describe a statistical method for analysis of data obtained by SAGE.

No abstract

In Saccharomyces cerevisiae, peroxisomes are the exclusive site for the degradation of fatty acids. Upon growth with the fatty acid oleic acid as sole carbon source, not only are the enzymes of beta‐oxidation and catalase A induced, but also the peroxisomal compartment as a whole increases in volume and the number of organelles per cell rises. We previously identified a cis‐acting DNA sequence [oleate response element (ORE)] involved in induction of genes encoding peroxisomal proteins. The aim of our investigation was to test whether a single mechanism acting via the ORE coordinates the events necessary for the proliferation of an entire organelle. Here we report the cloning and characterization of the oleate‐specific transcriptional activator protein Pip2p (pip: peroxisome induction pathway). Pip2p contains a typical Zn(2)‐Cys(6) cluster domain and binds to OREs. A pip2 deletion strain is impaired in growth on oleate as sole carbon source and the induction of beta‐oxidation enzymes is abolished. Moreover, only a few, small peroxisomes per cell can be detected. These results indicate that fatty acids activate Pip2p, which in turn activates the transcription of genes encoding beta‐oxidation components and acts as the crucial activator of peroxisomes.

In the yeast Saccharornyces cerevisiae, two transcriptional activators belonging to the Zn,Cys, protein family, Pip2p and Oaflp, are involved in fatty-acid-dependent induction of genes encoding peroxisomal proteins. This induction is mediated via an upstream activation sequence called the oleate-response element (ORE). DNA-bandshift experiments with ORE probes and epitope-tagged proteins showed that two binary complexes occurred: in wild-type cells the major complex consisted of a Pip2p . Oaflp heterodimer, but in cells in which Oaflp was overexpressed an Oaflp homodimer was also observed. The genes encoding Oaflp and Pip2p were controlled in different ways. The OAFl gene was constitutively expressed, while the PIP2 gene was induced upon growth on oleate, giving rise to positive autoregulatory control. We have shown that the Pip2p . Oaflp heterodimer is responsible for the strong expression of the genes encoding peroxisomal proteins upon growth on oleate. Pip2p and Oaflp form an example of a heterodimere of yeast Zn,Cys, zinc-finger proteins binding to DNA.