Dynamics of gene expression revealed by comparison of serial analysis of gene expression transcript profiles from yeast grown on two different carbon sources

Abstract

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 (>20%) 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 approximately 6300 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 beta-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.

Figure 1

Display of steady-state mRNA levels in yeast grown on different carbon sources. Each bar represents one mRNA species. (A) Display of mRNA levels in glucose-grown wild-type yeast. The 500 highest expressed genes are sorted by levels of mRNA copies per cell. (B) mRNA levels in oleate-grown wild-type cells. The genes are sorted in the same order as in A. (C) mRNA levels of the 500 highest expressed genes in oleate-grown wild-type cells. The genes are sorted by levels of mRNA copies per cell. (D) mRNA levels in oleate-grown pip2/oaf1 cells. The genes are sorted in the same order as in C.

Figure 2

mRNA levels of genes encoding peroxins and peroxisomal matrix enzymes. (A) Bars in the front row represent mRNA levels in glucose-grown wild-type cells (wt glu); bars in the back row represent mRNA levels in oleate-grown wild-type cells (wt ole). Note that all peroxins localize to the left, whereas all matrix enzymes localize to the right, stressing the differences in expression levels between the two classes of genes. (B). Bars in the back row represent mRNA levels in oleate-grown wild-type cells (wt ole); bars in the front row represent mRNA levels in oleate-grown pip2/oaf1 cells (p/o ole). *, Statistically significant difference in expression levels (p < 0.05).

Figure 3

mRNA levels of genes encoding mitochondrial proteins. The 100 highest expressed genes in wild-type cells grown in oleate medium are displayed in the back row (wt ole) and are sorted by numbers of mRNA copies per cell. The front row represents the mRNA levels for the same genes of wild-type cells grown on glucose (wt glu).

Figure 4

Transport of reducing equivalents among peroxisomes, cytoplasm, and mitochondria requires membrane-localized metabolite transporters. Indicated are proposed shuttles for malate/oxaloacetate, isocitrate/2-oxoglutarate, and glycerol-3-phosphate/dihydroxyacetonephosphate. The question mark indicates a cytosolic glycerol-3-phosphate dehydrogenase, of which the activity was measured in the cytosol (van Roermund, Hettema, and Tabak, unpublished observations).

Figure 5

mRNA levels of genes encoding proteins for gluconeogenesis, glyoxylate cycle, and heat shock and stress response. (A) Bars in the front row represent mRNA levels in glucose-grown wild-type cells (wt glu); bars in the back row represent mRNA levels in oleate-grown wild-type cells (wt ole). (B). Bars in the front row represent mRNA levels in oleate grown pip2/oaf1 cells (p/o ole); bars in the back row represent mRNA levels in oleate grown wild-type cells (wt ole). *, Statistically significant difference in expression levels (p < 0.05).

Figure 6

Statistical analysis. (A) Ninety-five percent confidence intervals (thin lines) for the number of copies per cell found in the oleate condition (thick line) and critical values for the statistical comparison of these numbers with numbers found in a control condition. Control values that fall outside the critical values can be considered different from the corresponding control value at significance levels of 0.05 (▴), 0.01 (♦) and 0.001 (▪), respectively. Calculations are carried out for a sample size of 10,366 sequenced tags in the oleate condition and 60,633 in the control condition, assuming the total number of mRNA molecules in a yeast cell to be 15,000. (B). Number of tags required in the experimental group to detect a 2- to 20-fold induction in abundance of mRNA compared with the control group from which 60,633 tags already have been sequenced. Calculations were performed with the SAGEstat software, assuming the total number of mRNA molecules in a yeast cell to be 15,000, with α = 0.05 and β = 0.1.