Distinct classes of sporulation-specific genes are sequentially expressed during the process of spore formation in Saccharomyces cerevisiae. The transition from expression of early meiotic genes to expression of middle sporulation-specific genes occurs at about the time that cells exit from pachytene and form the meiosis I spindle. To identify genes encoding potential regulators of middle sporulation-specific gene expression, we screened for mutants that expressed early meiotic genes but failed to express middle sporulation-specific genes. We identified mutant alleles of RPD3, SIN3, and NDT80 in this screen. Rpd3p, a histone deacetylase, and Sin3p are global modulators of gene expression. Ndt80p promotes entry into the meiotic divisions. We found that entry into the meiotic divisions was not required for activation of middle sporulation genes; these genes were efficiently expressed in a clb1 clb3 clb4 strain, which fails to enter the meiotic divisions due to reduced Clbdependent activation of Cdc28p kinase. In contrast, middle sporulation genes were not expressed in a dmc1 strain, which fails to enter the meiotic divisions because a defect in meiotic recombination leads to a RAD17-dependent checkpoint arrest. Expression of middle sporulation genes, as well as entry into the meiotic divisions, was restored to a dmc1 strain by mutation of RAD17. Our studies also revealed that NDT80 was a temporally distinct, pre-middle sporulation gene and that its expression was reduced, but not abolished, on mutation of DMC1, RPD3, SIN3, or NDT80 itself. In summary, our data indicate that Ndt80p is required for expression of middle sporulation genes and that the activity of Ndt80p is controlled by the meiotic recombination checkpoint. Thus, middle genes are expressed only on completion of meiotic recombination and subsequent generation of an active form of Ndt80p.Sporulation is a process of cellular differentiation that is initiated in diploid a/␣ cells of the yeast Saccharomyces cerevisiae in response to nitrogen starvation in the presence of a nonfermentable carbon source. As a cell progresses through the events of meiosis and spore wall formation, a coordinated series of genetic and morphological events generates a tetrad of haploid spores within an ascus (reviewed in references 9 and 30). The sporulation program begins when starved cells exit the mitotic cell cycle and undergo premeiotic DNA replication. This is followed by a lengthy prophase in which homologous chromosomes pair, the synaptonemal complex (SC) is elaborated, and a high level of genetic recombination occurs. At pachytene, the penultimate stage of prophase and the last stage at which cells can return to mitotic growth, the chromosomes are fully synapsed, and the spindle pole bodies lie sideby-side in the nuclear envelope. As cells exit from pachytene, the SC disassembles, chiasmata appear, and the spindle pole bodies separate to form the meiosis I spindle. The two meiotic divisions occur in rapid succession, leading first to segregation of homologous chromosome...

During sporulation of Saccharomyces cerevisiae, the four haploid nuclei generated by meiosis are encapsulated within multilayered spore walls. Taking advantage of the natural fluorescence imparted to yeast spores by the presence of a dityrosine-containing macromolecule in the spore wall, we identified and cloned two genes, termed DIT1 and DIT2, which are required for spore wall maturation. Mutation of these genes has no effect on the efficiency of spore formation or spore viability. The mutant spores, however, fail to accumulate the spore wall-specific dityrosine and lack the outermost layer of the spore wall. The absence of this cross-linked surface layer reduces the resistance of the spores to lyric enzymes, to ether, and to elevated temperature. Expression of the DIT and DIT2 genes is restricted to sporulating cells, with the DIT1 transcripts accumulating at the time of prospore enclosure and just prior to the time of dityrosine biosynthesis. Both genes act in a sporeautonomous manner implying that at least some of the activities responsible for forming the outermost layer of the spore wall reside within the developing spore rather than in the surrounding ascal cytoplasm. As the DIT2 gene product has significant homology with cytochrome P-450s, DIT2 may be responsible for catalyzing the oxidation of tyrosine residues in the formation of dityrosine.

The NDT80 gene of Saccharomyces cerevisiae, which encodes a global activator of transcription of middle sporulation-specific genes, is first expressed after the activation of early meiotic genes but prior to activation of middle sporulation-specific genes. Both upstream repression sequence 1 (URS1) and mid-sporulation element (MSE) sites are present in the promoter region of the NDT80 gene; these elements have been shown previously to contribute to the regulation of expression of early and middle sporulation-specific genes, respectively, by mediating repression in growing cells and activation at specific times during sporulation. In this study, we have shown that the overlapping windows of URS1-and MSE-mediated repression and activation are responsible for the distinctive premiddle expression pattern of the NDT80 gene. Our data suggest that a Sum1-associated repression complex bound at the NDT80 MSE sites prevents Ime1 tethered at the NDT80 URS1 sites from activating transcription of the NDT80 gene at the time that Ime1-dependent activation of early URS1-regulated meiotic genes is occurring. We propose that a decrease in the efficiency of Sum1-mediated repression as cells progress through the early events of the sporulation program allows the previously inactive Ime1 tethered at the URS1 NDT80 sites to promote a low level of expression of the NDT80 gene. This initial phase of URS1-dependent NDT80 expression is followed by Ndt80-dependent upregulation of its own expression, which requires the MSE NDT80 sites and occurs concomitantly with Ndt80-dependent activation of a set of middle MSE-regulated sporulation-specific genes. Mutation of IME2 prevents expression of NDT80 in sporulating cells. We show in this study that NDT80 is expressed and that middle genes are activated in cells of an ⌬ime2/⌬ime2 ⌬sum1/⌬sum1 strain in sporulation medium. This suggests that Ime2 activates expression of NDT80 by eliminating Sum1-mediated repression.The sporulation program of the yeast Saccharomyces cerevisiae provides a simple model system to study the temporal control of gene expression during development. On entry into the sporulation program, which is triggered by starvation, a diploid a/␣ cell completes one round of premeiotic DNA replication and then progresses through a lengthy prophase during which a high level of recombination occurs and homologous chromosomes pair. Cells then undergo the reductional and equational meiotic divisions, which result in a single four-lobed nucleus that contains the four haploid complements of chromosomes. The nuclear lobes are engulfed and ultimately pinched off by the prospore membranes that extend from the spindle pole bodies. The deposition of spore wall material within the prospore membrane completes the sporulation process, generating four mature spores arranged tetrahedrally within the ascus.Studies aimed at identifying genes that are differentially expressed as cells progress through sporulation or that serve sporulation-specific roles have defined four temporally distinct classes of sp...

During spoiulation of Saccharomyces cerevisiae, meiosis is followed by encapsulation of haploid nuclei within multilayered spore walls. Completion of the late events of the sporulation program requires the SPSl gene. This developmentally regulated gene, which is expressed as cells are nearing the end of meiosis, encodes a protein with homology to serine/threonine protein kinases. The catalytic domain of Spsl is 44% identical to the kinase domain of yeast Ste20, a protein involved in the pheromone-induced signal transduction pathway. Cells of a MATa./MATa. spsl/spsl strain arrest after meiosis and fail to activate genes that are normally expressed at a late time of sporulation. The mutant cells do not form refractile spores as assessed by phase-contrast microscopy and do not display the natural fluorescence and ether resistance that is characteristic of mature spores. Examination by electron microscopy reveals, however, that prospore-like compartments form in some of the mutant cells. These immature spores lack the cross-linked surface layer that surrounds wild-type spores and are more variable in size and number than are the spores of wild-type cells. Despite their inability to complete spore formation, spsl-arrested cells are able to resume mitotic growth on transfer to rich medium, generating haploid progeny. Our results suggest that the developmentally regulated Spsl kinase is required for normal progression of transcriptional, biochemical, and morphological events during the later portion of the sporulation program.