Identification of sequences responsible for transcriptional activation of the allantoate permease gene in Saccharomyces cerevisiae.

Abstract

DAL5 is a constitutively expressed allantoin system gene whose product is required for allantoate transport. Its simple pattern of expression prompted us to use this gene for identifying the element(s) that mediates transcriptional activation of allantoin system genes. Deletion analysis of the DAL5 5'-flanking sequences resulted in identification of two small regions required for DAL5 expression. Analysis of these two regions with synthetic oligonucleotides localized the sequences supporting transcriptional activation to two DNA fragments of 10 to 12 base pairs, each containing one copy of the pentanucleotide 5'-GATAA-3'. The 5'-flanking region of DAL5 contained eight copies of this sequence. Synthetic constructions containing single copies of these fragments were unable to support transcriptional activation, while those containing two or more copies supported high-level activation. The 5'-GATAA-3' sequence was also found beneath the footprint of a DNA-binding protein. These observations are consistent with the suggestion that DNA fragments containing the sequence 5'-GATAA-3' play an important role in DAL5 gene expression, probably representing a portion of the binding site for a transcriptional activation factor.

Nitrogen starvation induces expression of Lg‐FLO1 and flocculation in bottom‐fermenting yeast

“…Lg-FLO1 expression increased 3.0-fold in nitrogen-starved cells. As previously reported, DAL5 transcription was induced under nitrogen starvation conditions (Rai et al, 1989) and was also induced in bottom-fermenting yeast under nitrogen starvation conditions (Figure 3). lacZ activity in the laboratory yeast YPH500 transformed with pYABE-LgFLO1 (P Lg-FLO1 -lacZ) is shown in The flocculation of bottom-fermenting yeast is controlled by a mechanism linked to nitrogen catabolite repression…”

Mks1p Is Required for Negative Regulation of Retrograde Gene Expression in Saccharomyces cerevisiae but Does Not Affect Nitrogen Catabolite Repression-sensitive Gene Expression

“…Uracil (20 mg/liter), leucine (120 mg/liter), and tryptophan (20 mg/liter) were added to provide for auxotrophic requirements. ␤-galactosidase assays were performed as described (5).…”

Functional Domain Mapping and Subcellular Distribution of Dal82p in Saccharomyces cerevisiae

“…EMSAs were performed using 10 g of calf thymus ϩ 50 ng of radiolabeled DNA fragment and 1-5 g of protein (crude cell extract) (49). Urea amidolyase activity was determined in nystatin-permeabilized cells/transformants (4,50,51).…”

“…The first and least complex of these mechanisms is exemplified by the allantoin permease gene (DAL5), which is subject only to nitrogen catabolite repression (NCR) 1 (1)(2)(3)(4)(5). NCR is the physiological process through which the expression of genes encoding proteins needed for the transport and catabolism of poor nitrogen sources is maintained at low levels when a good nitrogen source is available (6,7).…”