Bipartite structure of an early meiotic upstream activation sequence from Saccharomyces cerevisiae.

Diploid a/alpha Saccharomyces cerevisiae cells cease mitotic growth and enter meiosis in response to starvation. Expression of meiotic genes depends on the IME1 gene product, which accumulates only in meiotic cells. We report here an analysis of the regulatory region of IME2, an IME1-dependent meiotic gene. Deletion and substitution studies identified a 48-bp IME1-dependent upstream activation sequence (UAS). Activity of the UAS also requires the RIM11, RIM15, and RIM16 gene products, which are required for expression of the chromosomal IME2 promoter and for meiosis. Through a selection for suppressors that permit UAS activity in an ime1 deletion mutant, we identified recessive mutations in three genes, SIN3 (also called RPD1, UME4, and SDI1), RPD3, and UME6 (also called CAR80), that were previously known as negative regulators of other early meiotic genes. Mutational analysis of the IME2 UAS reveals two critical sequence elements: a G+C-rich sequence (called URS1), previously identified at many meiotic genes, and a newly described element, the T4C site, that we found at a subset of meiotic genes. In agreement with prior studies, URS1 mutations lead to elevated IME2 UAS activity in the absence of IME1. However, the URS1 mutations prevent any further stimulation of UAS activity by IME1. Repression through URS1 has been shown to require the UME6 gene product. We find that activation of the IME2 UAS by IME1 also requires the UME6 gene product. Thus, UME6 and the URS1 site both have dual negative and positive roles at the IME2 UAS. We propose that IME1 modifies UME6 to convert it from a negulator to a positive Regulor.     

The DAL7 promoter consists of multiple elements that cooperatively mediate regulation of the gene''s expression.

Expression of the allantoin system genes in Saccharomyces cerevisiae is induced by allophanate or its analog, oxalurate. This work provides evidence for the involvement of distinct types of cis-acting elements in the induction process. The first element was found to have the properties of an upstream activation sequence (UAS). This element was localized to a 16-base-pair (bp) DNA fragment containing a short 5-bp sequence that occurred repeatedly in the upstream region of DAL7. When present in two or more copies, the 16-bp fragment supported high-level beta-galactosidase production in a CYC1-lacZ expression vector; there was, however, no response to the allantoin pathway inducer. The second element had the properties of a negatively acting element or upstream repression sequence (URS). This element was localized to a 16-bp DNA fragment containing an 8-bp sequence that was repeated four times in the upstream region of DAL7. A fragment containing the 8-bp repeated sequence placed adjacent to the UAS-containing fragment mediated inhibition of the ability of the UAS to support lacZ expression regardless of whether inducer was present. A third element, designated an upstream induction sequence (UIS), was required for response to inducer. The UIS was localized to a small DNA fragment containing an approximately 10-bp sequence that was repeated twice in the upstream region of DAL7. When a fragment containing the 10-bp repeated sequence was placed adjacent to these UAS and URS elements, the construction (UIS-UAS-URS) supported normal oxalurate-mediated induction of beta-galactosidase synthesis. These data are consistent with the suggestion that multiple, cis-acting elements participate in the induction process.     

Functional analysis of the regulatory region of the yeast phosphatidylserine synthase gene, PSS.

The upstream region of the PSS gene contains three positive cis-acting elements, upstream activation sequences 1 and 2 (UAS1 and UAS2) and a TATA box. The 5' end of UAS1 occurs between positions -239 and -209, and that of UAS2 is between positions -172 and -164. UAS2 contains 5'-TTCACATG-3' as a core sequence at positions -161 to -154. Mutational analysis revealed that this octamer is responsible for the control of PSS expression by inositol and choline. The TATA box is located at positions -112 to -108. In addition, PSS contains a negative cis-acting sequence between UAS2 and the TATA box.     

Stimulation of later functions of the yeast meiotic protein kinase Ime2p by the IDS2 gene product.

Ime2p is a protein kinase that is expressed only during meiosis in Saccharomyces cerevisiae. Ime2p stimulates early, middle, and late meiotic gene expression and down-regulates expression of IME1, which specifies an activator of early meiotic genes that acts independently of Ime2p. We have identified a new gene, IDS2 (for IME2-dependent signaling), which has a functional relationship to Ime2p. An ids2 null mutation delays down-regulation of IME1 and expression of middle and late meiotic genes. In an ime1 null mutant that express IME2 from the GAL1 promoter (ime1 delta PGAL1-IME2 mutant), early meiotic gene expression depends only upon Ime2p. In such strains, Ids2p is dispensable for expression of the early genes HOP1 and SPO13 but is essential for expression of the middle and late genes SPS1, SPS2, and SPS100. Ids2p is also essential for the autoregulatory pathway through which Ime2p activates its own expression via the IME2 upstream activation sequences (UAS). An PGAL1-IME2 derivative that produces a truncated Ime2p (lacking its C-terminal 174 residues) permits IME2 UAS activation in the absence of Ids2p. This observation suggests that Ids2p acts upstream of Ime2p or that Ids2p and Ime2p act in independent, convergent pathways to stimulate IME2 UAS activity. Accumulation of epitope-tagged Ids2p derivatives is greatest in growing cells and declines during meiosis. We propose that Ids2p acts indirectly to modify Ime2p activity, thus permitting Ime2p to carry out later meiotic functions.