Analysis of the Upstream Regulatory Region of the GTS1 Gene Required for Its Oscillatory Expression

Abstract

The protein level of the GTS1 gene product (Gts1p) fluctuated during the oscillation of energy metabolism in continuous culture of the yeast Saccharomyces cerevisiae. Here, we found that the GTS1 mRNA level oscillated with the same periodicity as the metabolic oscillation, suggesting that the expression of GTS1 was regulated at the transcriptional level. As the 5'-upstream sequence of GTS1 contains two short open-reading frames at -310 and -829 bp from the initiation codon, we determined the GTS1 promoter required for the oscillatory expression. The upstream sequence was truncated into fragments of 183, 355, 1,042, and 1,572 bp, named GTS1pr.183 and so on, and their effects on the expression of lacZ as a reporter gene and the GTS1 gene itself were examined. The beta-galactosidase activity and Gts1p level oscillated in the continuous cultures when genes were expressed under the control of GTS1pr.183 but not GTS1pr.355. The disappearance of the metabolic and cell-cycle oscillations in the GTS1-deleted mutant was rescued by the transformation with GTS1pr.183-GTS1 but not with GTS1pr.355-GTS1. However, the stress-resistance oscillations were not found in the cells transformed with GTS1pr.183-GTS1, differing from the case of GTS1pr.1042-GTS1 reported previously [Wang et al. (2001) FEBS Lett. 489, 81-86]. Thus, we suggest that the 183-bp upstream sequence of GTS1 is basically required for the metabolic oscillation, while the 1,042-bp upstream sequence is required for oscillations of stress resistance.