| The biosynthetic pathway leading to the formation of ergosterol in yeast has been well conserved making this simple eukaryotic organism an excellent model system in which to study sterol biosynthesis and regulation. In this investigation, three ergosterol biosynthetic genes, from the terminal portion of the pathway, were characterized based on DNA sequence analysis, gene disruption, and in the case of ERG3, regulation of expression. ERG2, ERG3, and ERG5 have been shown to be non-essential for the aerobic viability of Saccharomyces cerevisiae. The sterol molecules which accumulate in mutant cells blocked at any one of these steps are capable of functionally substituting for ergosterol within the plasma membrane. All three gene products are localized to microsomes in yeast. Based on the hydrophobic nature of each protein, as well as the presence of potential endoplasmic reticulum retrieval signals in the ERG2 and ERG3 amino acid sequences, these enzymes are likely endoplasmic reticulum-membrane bound proteins. Analysis of the ERG5 sequence revealed the presence of a cytochrome P450 signature motif. ERG5 is the second cytochrome P450 involved in ergosterol biosynthesis. Analysis of ERG3 expression in response to various physiological signals indicates that oxygen and heme are positive regulators of ERG3 transcription, and their effect is mediated through the action of the HAP1 trans-activating factor. Finally, these studies demonstrate that ERG3 is subject to feedback regulation by ergosterol in a manner similar to that observed for cholesterol biosynthesis in animal cells. |
