Genes encoding adenine biosynthetic enzymes in Saccharomyces cerevisiae, including ADE5,7, ADE8, ADE4, ADE1, and ADE2, are coordinately repressed at the transcriptional level when adenine is added to the growth medium (11,33,34,44). The expression of HIS4, encoding three activities in the histidine pathway, is similarly regulated by exogenous adenine (45). Bas1p and Bas2p are DNA binding proteins required for transcription of HIS4 (3) and ADE genes (11, 44) in medium lacking adenine. Bas1p binds to DNA through an N-terminal Myb-related domain (23) to sequences containing 5Ј-TGAC TC-3Ј as a common core element (3,11,45). Binding sites for Bas1p were identified in the HIS4 (3, 45), ADE2, and ADE5,7 promoters in vitro and shown to be required for transcription of the corresponding genes in adenine-free medium (11, 34). Bas2p is a homeodomain protein (9) that binds in vitro to an AϩT-rich sequence in the HIS4 promoter located adjacent to the Bas1p binding site that is required for efficient HIS4 transcription in vivo (3, 45). It was also shown that Bas2p binds to an ADE2 promoter fragment containing the critical Bas1p binding site; however, the exact location of the Bas2p binding site(s) at ADE2 was not determined (11).Based on the fact that mutations in BAS1 or BAS2 lead to low, unregulated expression of ADE genes (11, 44) and HIS4 (3, 45), it was proposed that adenine repression occurs by reducing the ability of Bas1p or Bas2p to stimulate transcription of these genes. In accordance with this idea, we recently identified a 74-bp fragment from ADE5,7 containing two Bas1p binding sites, an Abf1p binding site, and a Bas2p binding site that is sufficient to confer adenine-regulated transcription on the heterologous CYC1 promoter (Ade upstream activating sequence [UAS Ade ] function). Mutations in the Abf1p, Bas1p, or Bas2p sites that impaired UAS Ade function in vivo abolished or weakened binding of the corresponding proteins to the ADE5,7 sequences in vitro. It is noteworthy that none of ca. 60 substitution mutations in UAS Ade produced the constitutively derepressed phenotype expected from inactivation of a negative control site (34). These results are consistent with the idea that adenine repression occurs by reducing the expression or function of Bas1p, Bas2p, or Abf1p, rather than through binding of a repressor protein to the promoter. Because addition of adenine to the medium did not affect expression of BAS1-lacZ (45) or BAS2 (11), the ability of Bas1p or Bas2p to activate transcription, rather than their expression, may be down-regulated by adenine.Among the three positive regulators implicated at ADE5,7, we considered Bas1p to be the most likely target for negative regulation by adenine. Abf1p and Bas2p both function at other promoters that presumably are unresponsive to purine levels. Abf1p was first identified by its binding to autonomous replicating sequences (12) and subsequently for its role in the silencing of HML and HMR (8). In addition, Abf1p binds to and activates the promoters of many genes involve...
The analysis of cell fate patterning during the vulval development of Caenorhabditis elegans has relied mostly on the direct observation of cell divisions and cell movements (cell lineage analysis). However, reconstruction of the developing vulva from EM serial sections has suggested seven different cell types (vulA, vulB1, vulB2, vulC, vulD, vulE, and vulF), many of which cannot be distinguished based on such observations. Here we report the vulval expression of seven genes, egl-17, cdh-3, ceh-2, zmp-1, B0034.1, T04B2.6 and F47B8.6 based on gfp, cfp and yfp (green fluorescent protein and color variants) reporter fusions. Each gene expresses in a specific subset of vulval cells, and is therefore useful as a marker for vulval cell fates. Together, expressions of markers distinguish six cell types, and reveal a strict temporal control of gene expression in the developing vulva.
The great-grandprogeny of the Caenorhabditis elegans vulval precursor cells (VPCs) adopt one of the final vulA, B1, B2, C, D, E, and F cell fates in a precise spatial pattern. This pattern of vulval cell types is likely to depend on the cis-regulatory regions of the transcriptional targets of intercellular signals in vulval development. egl-17, zmp-1, and cdh-3 are expressed differentially in the developing vulva cells, providing a potential readout for different signaling pathways. To understand how such pathways interact to specify unique vulval cell types in a precise pattern, we have identified cis-regulatory regions sufficient to confer vulval cell type-specific regulation when fused in cis to the basal pes-10 promoter. We have identified the C. briggsae homologs of these three genes, with their corresponding control regions, and tested these regions in both C. elegans and C. briggsae. These regions of similarity in C. elegans and C. briggsae upstream of egl-17, zmp-1, and cdh-3 promote expression in vulval cells and the anchor cell (AC). By using the cis-regulatory analysis and phylogenetic footprinting, we have identified overrepresented sequences involved in conferring vulval and AC expression.
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