We have previously shown that transcript levels expressed from the yeast TMPI gene fluctuate periodically during the yeast cell cycle. However, it was not known whether periodic expression resulted from a regulatory mechanism acting at the level of transcription or a regulatory mechanism acting at the level of cell cycle stage-dependent changes in the stability of the TMPI transcript. In this report we now show that the periodic expression of TMPI transcript is primarily controlled at the level of its transcription by sequences which are upstream of its transcription initiation sites. We also localized the upstream sequences necessary for periodic transcription to a 150-base-pair region and show that this region encodes an element(s) with the properties of a periodic upstream activating sequence. The regulatory region defined in this study apparently does not contain consensus sequences similar to those reported for the cell cycle-regulated HO endonuclease or for the histone H2A and H2B genes of Saccharomyces cerevisiae.One approach to investigating the processes which control eucaryotic cell division is to identify and study those genes which are differentially regulated during the cell cycle. If progression through the cell division cycle results from the sequential transient expression of dormant genes, then identifying the factors which govern the expression of these genes is of considerable interest. In the yeast Saccharomyces cerevisiae, only a few genes are currently known to exhibit differential expression during the cell cycle. These include the thymidylate synthase gene TMPJ (25), the histone H2A and H2B genes of the TRTJ locus (12), the HO endonuclease gene (20), the thymidylate kinase gene CDC8 (28), and the DNA ligase gene CDC9 (23).Previous studies have shown that the regulation of the periodically expressed yeast histone genes results from complex controls acting at both the transcriptional and posttranscriptional levels (14,21,22). In addition, their proper expression may also involve the placement of these genes adjacent to an origin of DNA replication (21). Similarly, expression of the HO gene appears to be governed by complex controls acting transcriptionally and posttranscriptionally (4,20). Nasmyth (20) has identified a 12-base-pair (bp) consensus sequence which is repeated many times within the URS2 region of HO and has been shown to be critically involved in the periodic expression of this gene.We found that TMPJ mRNA is transiently expressed during the cell cycle, with peak amounts occurring during late G1 and early S phase just after the cdc28la-factor arrest point (25). In a recent study, White et al. (28) demonstrated that the periodic expression of TMPJ, CDC8, and CDC9 mRNA was coincident, while the histone H2A gene was expressed distinctly later in the cell cycle. This was shown by analyzing the times at which these genes were expressed after release from G1 arrest and also by showing that only the periodicity of the histone genes was influenced by the cdc4-3 mutation. These results ...