Upstream activating sequence

About: Upstream activating sequence is a research topic. Over the lifetime, 1633 publications have been published within this topic receiving 100112 citations.

Chromatin structure and regulation of the eukaryotic regulatory gene GAL80

Abstract: The chromatin structure around the 5' end of the yeast regulatory gene GAL80 has been determined. The chromatin organization is very similar to that on the 5' regions of the GAL1-10 structural genes: a constitutive hypersensitive region containing the upstream activating sequence (UAS) element, and nucleosomes around this hypersensitive region. The downstream nucleosome, which is a positioned nucleosome, covers the TATA and transcription start sites. The nucleosome upstream of the hypersensitive region undergoes significant change when cells are grown in galactose, where GAL80 gene expression is induced to maximal levels. The change may be related to the induction process. GAL4 protein binds strongly to the GAL80 UAS in galactose-grown cells, less strongly in glycerol-grown cells, and not at all in glucose-grown cells. These data and published gene expression data are used to develop a model for the regulation of the GAL80 regulatory gene.

Characterization of the Rat mdr2 Promoter and Its Regulation by the Transcription Factor Sp1

Abstract: The mdr2 gene encodes a P-glycoprotein that transports phospholipids across the canalicular membrane in hepatocytes. In this report we describe the isolation, sequencing and first functional characterization of the promoter of mdr2. Analysis of 1.6 kb of DNA upstream of the initiation of translation revealed that this sequence has a high GC content, lacks a TATA element and contains a number of putative transcription factor binding sites. We observed that transcription initiates at several sites between -290 and -463 and that this region was critical for promoter activity. Gel mobility shift assays indicated that Sp1 protein binds to a Sp1 consensus site located at -263. Co-expression of Sp1 protein with a reporter construct containing the -263 GC box demonstrated that Sp1 regulates transcription of this promoter. Expression of a non-functional Sp1 protein did not increase transcription from the mdr2 promoter. Mutation of the -263 GC box diminished the response of the promoter to Sp1 protein. Mutation of this site also decreased expression of this promoter in cells which normally express this gene. These data show that Spl has a role in the regulation of mdr2 expression.

The nucleotide concentration determines the specificity of in vitro transcription activation by the sigma 54-dependent activator FhlA.

Abstract: An in vitro transcription system has been set up for formate- and FhlA-dependent transcription activation at the -12/-24 promoter of the fdhF gene from Escherichia coli by sigma 54-RNA polymerase. It requires the presence of the upstream activation sequence on supercoiled DNA. Transcription is independent from the effector formate at nucleoside triphosphate concentrations of 400 microM and above and completely dependent on the presence of the effector when the concentration is lowered to 300 microM. Inclusion of nucleoside diphosphates in the system raises the nucleoside triphosphate level at which specific induction by formate can take place. The threshold level of FhlA relative to that of template DNA required for transcription activation in the absence of formate was lowered at a high nucleoside triphosphate concentration. On the other hand, transcription activation at the fdhF promoter lacking the upstream activation sequence requires an increased ratio of FhlA to promoter plus the presence of formate; high ATP concentrations cannot bypass the effect of formate. These results are interpreted in terms of a model which implies that FhlA must undergo a change in its oligomeric state for transcription activation and that this oligomerization is favored by high nucleoside triphosphate concentrations, by the effector formate, and by the target DNA. In the absence of the target DNA, FhlA can line up at unspecific DNA and activate transcription; in this case, however, presence of formate and a higher FhlA concentration are required to stabilize and increase the amount of active oligomer.

Identification of engrailed promoter elements essential for interactions with a stripe enhancer in Drosophila embryos.

Abstract: Background The structures and functions of promoter sequences of most genes have been analysed using in vitro transcription and/or cultured cell systems, neither possessing tissue-specific enhancers. Promoter–enhancer interactions in vivo, in particular, during ontogeny, are still poorly understood. Results We have established a new method for the assessment of promoter activity in cells that participate in fly body formation, using the UAS/GAL4 system. A functional analysis was then conducted on the promoter sequence of the engrailed gene in Drosophila embryos. A 38-bp-long sequence, terminating with an initiator or RNA start site and a downstream promoter element, was found to be capable of receiving activation signals from the engrailed stripe enhancer. Transcriptional efficiency was improved significantly by the presence of upstream promoting elements, most functionally replaceable with synthetic GAGA factor binding sites. Conclusions We identified the in vivo minimum promoter of engrailed and demonstrated that the GAGA factor binding sites serve primarily as quantitative elements which augment transcriptional efficiency. Evidence was also obtained that indicated that not only enhancer but also promoter sequences were involved in the determination of the tissue-specificity of gene expression.