A regulatory region responsible for proline-specific induction of the yeast PUT2 gene is adjacent to its TATA box.

Abstract

Deletion analysis of the promoter of the PUT2 gene that functions in the proline utilization pathway of Saccharomyces cerevisiae identified a PUT2 upstream activation site (UAS). It is contained within a single 40-base-pair (bp) region located immediately upstream of the TATA box and is both necessary and sufficient for proline induction Transcriptional regulation of genes in the yeast Saccharomyces cerevisiae is mediated by the presence of various promoter elements, including upstream activation sites (UASs) (22,23,33,47,58,65), operator sites (34), repression or silencer sequences (7, 59), TATA boxes (24, 45, 52, 56), and initiation sites (24, 45). These elements are thought to be the binding sites of proteins that can transmit signals of change in the cellular environment to activate or repress expression (22,26,34,48,50) or the recognition sites for proteins of the transcriptional apparatus (57). The distance between the regulatory elements and the TATA box varies, and changes introduced in vitro often do not substantially affect gene function (3,15,28,31,36,56,57,60). Genes whose functions are related often share homologous upstream sequences that, in some cases, have been shown to be the binding sites for specific trans-acting regulators (2,22,29,30,48,50).Two genes whose expression is required for S. cerevisiae to use proline as a source of nitrogen are genetically unlinked and coordinately regulated at the transcriptional level. In the absence of preferred sources of nitrogen, the PUT] and PUT2 genes respond to induction by proline by an increase in the steady-state mRNA levels and a corresponding increase in proline oxidase and A'-pyrroline-5-carboxylate (PSC) dehydrogenase activities, respectively (8,11,37,61,62). The two enzymes carry signalling information that targets them to the mitochondrion, where they function to convert proline to glutamate (10, 62). Induction by proline requires a trans-acting positive regulator encoded by the PUT3 gene (8,9,12,61). Recessive, noninducible and semidominant, constitutive alleles of PUT3 have been isolated that affect the expression of both PUT] and PUT2 (9, 12). * Corresponding author.The aim of this study was to elucidate the promoter elements involved in proline induction of the PUT2 gene. A series of 5' and internal deletions constructed in vitro identified sequences essential for PUT2 promoter function.Our results indicate that this promoter has a rather simple structure, consisting of a single UAS located immediately adjacent to its TATA box. This UAS is essential for expression under both inducing and noninducing conditions and appears to be the site through which the PUT3 regulator acts. MATERIALS AND METHODSStrains and media. The S. cerevisiae strains used in this study were MB758-SB (MATa ura3-52), C74-4D (MATot ura3-52 PUT3-68), and C75-2B (MATa ura3-52 put3-75). The ura3-52 mutation was derived from strain DBY785 (laboratory of D. Botstein) and was introduced by crossing into isogenic derivatives of strain MB1000 (MATTa wild type [11]). The resulting...

Particle and genomic characteristics of a new member of the Ascoviridae: Diadromus pulchellus ascovirus.

“…Two types of transcription factor DNA-binding motif were found in the 494 bp repeats. The first corresponded to two overlapping motifs of ubiquitous factors, the mammalian NFI CS7 (position 336-350 ; Faisst & Meyer, 1992) and the fungal PUT2 UAS.1 (343-350 ; Siddiqui & Brandriss, 1988). The second type of motif corresponded to two well-conserved DNA binding sites (XGGAYGT) of the adenovirus E1A enhancer factor (Yoshida et al, 1989) …”

Symmetric cell division in pseudohyphae of the yeast Saccharomyces cerevisiae.

“…Yeast strains and transforming plasmids are listed in Tables 1 and 2. Strains used in these studies are isogenic to MB1000 (Brandriss and Magasanik, 1979), formerly 2:1278b (Grenson et al, 1966) and MB758-5B (Siddiqui and Brandriss, 1988). MB758-5B contains the ura3-52 mutation.…”

The Saccharomyces cerevisiae PUT3 activator protein associates with proline-specific upstream activation sequences.

“…gene under inducing and noninducing growth conditions. The PUT3 gene product was shown to act either directly or indirectly through this sequence (35).…”

“…This sequence, which is present once in the 819-bp PUT? promoter (35), is found twice in 458 bp of the PUT) promoter. Gel mobility shift assays demonstrated that the PUT UASs are the recognition sites of a DNA-binding complex that includes the PUT3 product.…”

“…A trans-acting positive regulator, the product of the PUT3 gene, mediates proline induction (6,9). PUT3 was defined genetically as a transcriptional activator based on the analysis of two mutant alleles that affected the levels of RNA accumulation and enzyme activity of the PUT genes (5,6,9,35,39). Promoter deletion analysis demonstrated the presence of a proline-specific UAS in a 40-base-pair (bp) sequence that was required for transcription of the PUT?…”