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PubMed1986

Sequences upstream of the STE6 gene required for its expression and regulation by the mating type locus in Saccharomyces cerevisiae.

The STE6 gene of Saccharomyces cerevisiae is an a-specific gene; it is repressed in alpha cells by the alpha 2 product of the mating type locus. To study the role of sequences upstream of STE6 in its regulation and expression, we have determined the DNA sequence of the promoter region, identified the start sites for the STE6 transcript, and identified sequences governing its transcription. Deletions that remove DNA upstream of the STE6 gene were produced and assayed for effects on regulation and expression. The deletions defined two intervals upstream of the STE6 transcription initiation sites. One contains all or part of a negative element; the other contains all or part of a positive element. The negative element is required for repression of STE6 by alpha 2: deletions lacking this element express STE6 constitutively. Such deletions remove a 31-base-pair site, located 135 base pairs upstream of the first transcript start site, that is highly homologous to sites present in the upstream regions of four other genes repressed by alpha 2. These sites are presumably responsible for repression of the a-specific genes by alpha 2. The positive element (a putative upstream activation site) is required for expression of STE6. The deletions define the left boundary of the proposed upstream activation site. Sequence homologies between STE6 and other a-specific genes are found in this region and may mediate activation of this set of genes.

Base Sequence↗

PubMed1999

Interaction between the skeletal muscle type 1 Na+ channel promoter E-box and an upstream repressor element. Release of repression by myogenin.

We have defined how four elements that regulate expression of the rat skeletal muscle type 1 sodium channel (SkM1) gene cooperate to yield specific expression in differentiated muscle. A basal promoter region containing within it a promoter E-box (-31/-26) is broadly expressed in many cells, including myoblasts and myotubes; mutations within the promoter E-box that disrupt binding of the myogenic basic helix-loop-helix (bHLH) factors reduce expression in all cell types only slightly. Sequential addition of upstream elements to the wild-type promoter confer increasing specificity of expression in differentiated cells, even though all three upstream elements, including a positive element (-85/-57), a repressor E-box (-90/-85), and upstream repressor sequences (-135/-95), bind ubiquitously expressed transcription factors. Mutations in the promoter E-box that disrupt the binding of the bHLH factors counteract the specificity conferred by addition of the upstream elements, with the greatest interaction observed between the upstream repressor sequences and the promoter E-box. Forced expression of myogenin in myoblasts releases repression exerted by the upstream repressor sequences in conjunction with the wild-type, but not mutant, promoter E-box, and also initiates expression of the endogenous SkM1 protein. Our data suggest that particular myogenic bHLH proteins bound at the promoter E-box control expression of SkM1 by releasing repression exerted by upstream repressor sequences in differentiated muscle cells.

Animals↗

PubMed1984

Stimulation of in vitro transcription by the upstream element of the adenovirus-2 major late promoter involves a specific factor.

We have previously reported that sequences located upstream from the TATA box of the Adenovirus-2 major late promoter (Ad2MLP), between -34 and -97, are necessary for efficient transcription in vivo and in vitro (1). We have utilized an in vitro competition assay to demonstrate that the upstream element requirement involves the binding of a specific factor(s), which results in stimulation of in vitro transcription from the Ad2MLP. DNA fragments prepared from Ad2MLP upstream sequence mutants which are transcribed much less efficiently than the wild-type promoter both in vivo and in vitro were shown to be unable to bind this factor. We have also constructed chimeric promoter recombinants containing the 21bp repeat upstream element of the SV40 early promoter inserted upstream from the Ad2MLP TATA box. The SV40 upstream element stimulates in vitro transcription from the heterologous Ad2MLP TATA box element; competition experiments show that the Ad2MLP upstream element-specific factor is different from the SV40-specific factor Sp1.

Adenoviruses, Human↗

PubMed1996

Multiprotein transcription factor UAF interacts with the upstream element of the yeast RNA polymerase I promoter and forms a stable preinitiation complex.

Like most eukaryotic rDNA promoters, the promoter for rDNA in Saccharomyces cerevisiae consists of two elements: a core element, which is essential, and an upstream element, which is not essential but is required for a high level of transcription. We have demonstrated that stimulation of transcription by the upstream element is mediated by a multiprotein transcription factor, UAF (upstream activation factor) which contains three proteins encoded by RRN5, RRN9, and RRN10 genes, respectively, and probably two additional uncharacterized proteins. The three genes were originally defined by mutants that show specific reduction in the transcription of rDNA. These genes were cloned and characterized. Epitope tagging of RRN5 (or RRN9), combined with immunoaffinity purification was used to purify UAF, which complemented all three (rrn5, rrn9, and rrn10) mutant extracts. Using rrn10 mutant extracts, a large stimulation by UAF was demonstrated for template containing both the core element and the upstream element but not for a template lacking the upstream element. In the absence of UAF, the mutant extracts showed the same weak transcriptional activity regardless of the presence or absence of the upstream element. We have also demonstrated that UAF alone makes a stable complex with the rDNA template, committing that template to transcription. Conversely, no such template commitment was observed with rrn10 extracts without UAF. By using a series of deletion templates, we have found that the region necessary for the stable binding of UAF corresponds roughly to the upstream element defined previously based on its ability to stimulate rDNA transcription. Differences between the yeast UAF and the previously studied metazoan UBF are discussed.

Amino Acid Sequence↗

PubMed1992

Role of the upstream region containing an intrinsic DNA curvature in the negative regulation of the phospholipase C gene of Clostridium perfringens.

The phospholipase C (alpha-toxin) gene (plc) of Clostridium perfringens was cloned into pUC19 and the effects of the upstream regions on expression of the plc gene were examined in Escherichia coli JM109. When the 0.7-kb region just upstream of the putative -35 site of the gene was deleted, production of phospholipase C increased approximately 10-fold. Northern blot hybridization analysis of the plc transcript showed that the upstream region inhibited transcription from the plc promoter. Nucleotide sequencing of this upstream region revealed that there are three periodically repeated (dA)5-6 tracts between positions -66 and -40 of the plc gene. A fragment containing this sequence showed anomalously slow electrophoretic mobility at low temperature, indicating that the region immediately upstream of the plc promoter is a locus of sequence directed DNA-bending. Nested deletions of the upstream region were created from its 5' end by exonuclease III and the effects of deletions on the expression of the plc gene were examined. When the 77-bp fragment containing the two (dA)5-6 tracts were deleted, phospholipase C production increased markedly. These results indicate that the intrinsic DNA curvature upstream of the plc promoter is involved in the negative regulation of the plc gene transcription.

Bacterial Toxins↗

PubMed1991

Analysis of the upstream activating sequence and site of carbon and nitrogen source repression in the promoter of an early-induced sporulation gene of Bacillus subtilis.

The transcription from the spoVG promoter of Bacillus subtilis is induced at the start of the stationary phase of growth and is dependent on the expression of the spoOA, spoOB, and spoOH genes. It is repressed in cells grown in the presence of excess glucose and glutamine and is under the negative control of the abrB gene. The spoOA and spoOB gene products function to suppress the negative control exerted by abrB. Transcription initiation requires the form of RNA polymerase holoenzyme that contains the spoOH gene product, sigma H. Optimal transcription also requires an upstream A-T-rich region termed the upstream activating sequence (UAS). The mechanism of UAS function was examined through mutational analysis of the spoVG promoter region. Deletion of the UAS or positioning the UAS one half turn or one full turn of the DNA helix upstream of its location in wild-type spoVG resulted in a severe reduction in promoter activity. Deletion of most of the UAS abolished the abrB-dependent repression of spoVG transcription. Higher activity was observed when the UAS was inserted 10 bp (one turn of the helix) upstream than when the sequence was repositioned either 5 or 13 bp upstream. Sequences upstream of the UAS were found not to be involved with the position-dependent function of the UAS. Positioning the UAS 42 or 116 bp upstream eliminated the stimulatory effect of the sequence on spoVG transcription. These data indicate that the UAS functions effectively when it is in close proximity to the -35 region. In vitro transcription analysis indicated that the deletion and insertion mutation affecting the UAS impair RNA polymerase-spoVG promoter interaction. Deletion of the UAS showed that the negative effect of exogenous glucose and glutamine is not dependent on the UAS but is exerted at a site within or near the -35 and -10 regions.

Bacillus subtilis↗

PubMed1998

Sequences just upstream of the simian immunodeficiency virus core enhancer allow efficient replication in the absence of NF-kappaB and Sp1 binding elements.

Large deletions of the upstream U3 sequences in the long terminal repeats (LTRs) of human immunodeficiency virus and simian immunodeficiency virus (SIV) accumulate in vivo in the absence of an intact nef gene. In the SIV U3 region, about 65 bp just upstream of the single NF-kappaB binding site always remained intact, and some evidence for a novel enhancer element in this region exists. We analyzed the transcriptional and replicative capacities of SIVmac239 mutants containing deletions or mutations in these upstream U3 sequences and/or the NF-kappaB and Sp1 binding sites. Even in the absence of 400 bp of upstream U3 sequences, the NF-kappaB site and all four Sp1 binding sites, the SIV promoter maintained about 15% of the wild-type LTR activity and was fully responsive to Tat activation in transient reporter assays. The effects of these deletions on virus production after transfection of COS-1 cells with full-length proviral constructs were much greater. Deletion of the upstream U3 sequences had no significant influence on viral replication when either the single NF-kappaB site or the Sp1 binding sites were intact. In contrast, the 26 bp of sequence located immediately upstream of the NF-kappaB site was essential for efficient replication when all core enhancer elements were deleted. A purine-rich site in this region binds specifically to the transcription factor Elf-1, a member of the ets proto-oncogene-encoded family. Our results indicate a high degree of functional redundancy in the SIVmac U3 region. Furthermore, we defined a novel regulatory element located immediately upstream of the NF-kappaB binding site that allows efficient viral replication in the absence of the entire core enhancer region.

Animals↗

PubMed2002

Deletion of Epstein-Barr virus regulatory sequences upstream of the EBNA gene promoter Wp1 is unfavorable for B-Cell immortalization.

Transcription of the six Epstein-Barr virus (EBV) EBNA genes is coordinately regulated, being driven by either the Cp promoter, which is encoded within the unique region just upstream of the EBV major internal repeat (IR-1), or by the Wp promoter, which is encoded within the IR-1 repeat and thus present in multiple copies. Previous analyses of Cp- and Wp-initiated transcription have identified a shared cis-regulatory element mapping to the region extending from -169 to -369 bp upstream of the Wp transcription initiation site (M. T. Puglielli, N. Desai, and S. H. Speck, J. Virol. 71:120-128, 1997). To assess the impact of this regulatory region on Cp and Wp activity in the context of the viral genome, we attempted to delete this regulatory region upstream of the first copy of Wp (Wp1). While 10 recombinant viruses were obtained in which this deletion was incorporated in the interior of the IR-1 repeat, only a single lymphoblastoid cell line (LCL) immortalized by a recombinant EBV harboring the deletion upstream of Wp1 was recovered. In contrast, using a control targeting vector in which the Wp regulatory sequences were intact but which contained a sequence tag within the W0 exon, we demonstrated that of the five recombinant viruses analyzed in which the crossover event had occurred upstream of the Wp sequence tag, four had incorporated the tagged sequences into Wp1 of the virus. Taken together, these results indicate that deletion of the regulatory sequences from -369 to -169 bp upstream of Wp1 is unfavorable for EBV-driven B-cell immortalization but is tolerated within the interior of the IR-1 repeat. Analysis of promoter usage in the clone 9-60 LCL, in which the W enhancer sequences were deleted upstream of Wp1, revealed the following: (i) the level of Cp-initiated transcription was significantly diminished compared to that of wild-type LCLs; (ii) the decreased Cp-initiated transcription was not efficiently compensated by transcription initiation from Wp1; and (iii) transcription initiation from downstream Wp promoters was detectable. This is the first report of an LCL in which transcription initiation from a Wp downstream of Wp1 has been documented.

B-Lymphocytes↗

PubMed1995

L-arginine-induced conducted signals alter upstream arteriolar responsivity to L-arginine.

Our purpose was to determine whether L-arginine was involved in vascular communication between downstream and upstream locations within a defined microvascular region. Arteriolar diameter was measured for the branches along a transverse arteriole in the superfused cremaster of anesthetized (pentobarbital sodium, 70 mg/kg i.p.) hamsters (N = 53). The upstream branch arterioles dilated significantly to locally applied L-arginine (100 mumol/L pipette concentration) only if the downstream branches (approximately 1400 microns away) were preexposed. With exposure order downstream to upstream, diameter change was last branch, -3.8 +/- 1.5% (of baseline); third, +58.1 +/- 27%; first, +92 +/- 26% (n = 5); with exposure order upstream to downstream: first branch, -0.4 +/- 3%; third, +5 +/- 11%; last, -5.6 +/- 7.5% (n = 4). Thus, downstream preexposure to L-arginine altered the responsivity upstream to locally applied L-arginine. Downstream-applied L-arginine also induced a conducted vasodilation (+17.8 +/- 2.8%; n = 14) 1327 +/- 166 microns upstream. This response was completely blocked by simultaneous sucrose (600 mOsm), halothane (0.0345%), or N omega-nitro-L-arginine (L-NNA, 100 mumol/L) exposure to the feed vessel (second micropipette) midway between the downstream site of L-arginine exposure and the upstream observation site. An acetylcholine-induced conducted vasodilation (+18.1 +/- 2.6%, n = 8) was also completely blocked by sucrose, halothane, or L-NNA.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗

PubMed2005

The seven E. coli ribosomal RNA operon upstream regulatory regions differ in structure and transcription factor binding efficiencies.

Ribosomal RNAs in E. coli are transcribed from seven operons, which are highly conserved in their organization and sequence. However, the upstream regulatory DNA regions differ considerably, suggesting differences in regulation. We have therefore analyzed the conformation of all seven DNA elements located upstream of the major E. coli rRNA P1 promoters. As judged by temperature-dependent gel electrophoresis with isolated DNA fragments comprising the individual P1 promoters and the complete upstream regulatory regions, all seven rRNA upstream sequences are intrinsically curved. The degree of intrinsic curvature was highest for the rrnB and rrnD fragments and less pronounced for the rrnA and rrnE operons. Comparison of the experimentally determined differences in curvature with programs for the prediction of DNA conformation revealed a generally high degree of conformity. Moreover, the analysis showed that the center of curvature is located at about the same position in all fragments. The different upstream regions were analyzed for their capacity to bind the transcription factors FIS and H-NS, which are known as antagonists in the regulation of rRNA synthesis. Gel retardation experiments revealed that both proteins interact with the upstream promoter regions of all seven rDNA fragments, with the affinities of the different DNA fragments for FIS and H-NS and the structure of the resulting complexes deviating considerably. FIS binding was non-cooperative, and at comparable protein concentrations the occupancy of the different DNA fragments varied between two and four binding sites. In contrast, H-NS was shown to bind cooperatively and intermediate states of occupancy could not be resolved for each fragment. The different gel electrophoretic mobilities of the individual DNA/protein complexes indicate variable structures and topologies of the upstream activating sequence regulatory complexes. Our results are highly suggestive of differential regulation of the individual rRNA operons.

Bacterial Proteins↗

PubMed2025

Translation of the downstream ORF from bicistronic mRNAs by human cells: Impact of codon usage and splicing in the upstream ORF.

Biochemistry textbooks describe eukaryotic mRNAs as monocistronic. However, increasing evidence reveals the widespread presence and translation of upstream open reading frames preceding the "main" ORF. DNA and RNA viruses infecting eukaryotes often produce polycistronic mRNAs and viruses have evolved multiple ways of manipulating the host's translation machinery. Here, we introduce an experimental model to study gene expression regulation from virus-like bicistronic mRNAs in human cells. The model consists of a short upstream ORF and a reporter downstream ORF encoding a fluorescent protein. We have engineered synonymous variants of the upstream ORF to explore large parameter space, including codon usage preferences, mRNA folding features, and splicing propensity. We show that human translation machinery can translate the downstream ORF from bicistronic mRNAs, albeit reporter protein levels are thousand times lower than those from the upstream ORF. Furthermore, synonymous recoding of the upstream ORF exclusively during elongation significantly influences its own translation efficiency, reveals cryptic splice signals, and modulates the probability of downstream ORF translation. Our results are consistent with a leaky scanning mechanism facilitating downstream ORF translation from bicistronic mRNAs in human cells, offering new insights into the role of upstream ORFs in translation regulation.

Humans↗

PubMed2006

Familial combined hyperlipidemia: upstream transcription factor 1 and beyond.

PURPOSE OF REVIEW: Familial combined hyperlipidemia is a common complex disease that accounts for up to 20% of premature coronary heart disease. The upstream transcription factor 1, located on 1q21, was recently shown to be linked and associated with familial combined hyperlipidemia in Finnish families. Upstream transcription factor 1 is the first gene identified by positional cloning for familial combined hyperlipidemia. Replication studies are critical to investigation of complex diseases because only they can verify the importance of the original findings. We review recent studies that examine the genetic contribution and functional consequence of upstream transcription factor 1 variants to familial combined hyperlipidemia and type 2 diabetes mellitus. Aiming beyond upstream transcription factor 1, we also evaluate novel strategies that have made it possible to globally examine the genome and the transcriptome. RECENT FINDINGS: Three independent studies support the role of upstream transcription factor 1 in familial combined hyperlipidemia. The results for type 2 diabetes mellitus and the metabolic syndrome have been less conclusive highlight novel strategies for gene identification in familial combined hyperlipidemia. SUMMARY: Currently, genetic and functional evidence is supportive of a role for upstream transcription factor 1 in the etiology of familial combined hyperlipidemia and its component traits, although the mechanism of causality still remains largely unknown.

Animals↗

PubMed1994

Cloning of the Syrian hamster p53 gene: structural and functional characterization of the upstream promoter region.

We isolated the p53 gene from Syrian hamster embryo cells by cosmid cloning procedures. The organization of the hamster p53 gene was similar to that of other mammalian p53 genes; it had 11 exons, a noncoding exon 1, and a long intron 1 (about 6.5 kb). The upstream p53 promoter was isolated, and the nucleotide sequence of a region encompassing 694 bp upstream from the exon/intron 1 boundary plus the first 6 nt of intron 1 was determined. This genomic region was highly homologous to those of mice and humans but contained a repetitive element not present in either species. Sequence comparisons with the murine and human promoters revealed the presence of similar transcription-factor binding motifs mapping within a 431-bp SacI-PstI fragment. Transient transfection assays of primary Syrian hamster embryo cells and neoplastic cell lines with recombinant constructs in which the SacI-PstI fragment was placed upstream of a bacterial chloramphenicol acetyltransferase (CAT) gene revealed the efficient expression of CAT activity. Primer extension analyses identified several putative transcription initiation sites within the p53 upstream promoter, the strongest of which was located 315 bp upstream from the exon/intron 1 junction and about 30 bp upstream from the region encompassing the regulatory motifs.

Animals↗

PubMed1993

Gene structure and upstream regulatory regions of human CYP2C9 and CYP2C18.

There is a genetic polymorphism in humans in the metabolism of S-mephenytoin which has been suggested to be mediated by either CYP2C18 or CYP2C9. We have isolated genomic clones for CYP2C9 and CYP2C18 from the liver of an individual phenotyped in vitro as an extensive metabolizer of S-mephenytoin. Analysis of the genes reveals nine coding exons spanning approximately 55 kb. The intron-exon organization was similar to that of other members of the CYP2C subfamily. Analysis of 2200 bp of 5' upstream sequence for CYP2C9 and 1300 bp 5' upstream sequence for CYP2C18 reveals canonical TATA boxes situated 57 bp upstream from the first codon, multiple consensus sequences for glucocorticoid regulatory elements, and identification of a 15 base sequence with high homology to a 5'-flanking sequence responsible for barbiturate-inducible expression of P450BM-3 in Bacillus megaterium. The upstream region for CYP2C9 was highly homologous (75%) to that of human CYP2C8 through most of the 2200 bp sequenced, but the upstream region of CYP2C18 was similar to CYP2C8 and CYP2C9 for only the first 200 bases. The availability of the sequences of the upstream regions and intron-exon junctions of CYP2C9 and CYP2C18 will allow future analysis of these genes in humans which differ in their ability to metabolize S-mephenytoin and other drugs.

Aryl Hydrocarbon Hydroxylases↗

PubMed1998

Role of far upstream repressor elements controlling proto-Ha-ras gene transcription.

The far upstream region of the rat Ha-ras gene has been characterized to determine whether possible repressor sequences may control the low level of Ha-ras gene transcription from its TATA-less, GC-rich strong promoter. The chloramphenicol acetyl transferase (CAT) gene under the control of the 3.8-kb Ha-ras upstream promoter was minimally expressed in HeLa cells. Surprisingly, CAT gene expression was increased by the deletion of a 0.7-kb BglII fragment containing non-coding exon minus 2 and TATA box promoter elements located 1.7 kb upstream of the GC-rich strong promoter. Far upstream (CA)25 repeats also appeared to repress Ha-ras gene activity. Sequences within the 0.7-kb BglII fragment suppressed CAT gene expression when placed upstream of a heterologous thymidine kinase (tk) gene promoter. Repressor activity was further localized to a 160-bp AvrII-BglII sub-fragment. Gel shift assays identified two sequence-specific DNA binding proteins. The results demonstrated for the first time that far upstream repressor sequences control normal transcription of the Ha-ras proto-oncogene.

Animals↗

PubMed1999

Isolation and functional characterization of the 5'-upstream region of mouse P/Q-type Ca2+ channel alpha1A subunit gene.

The omega-agatoxin-IVA-sensitive P/Q-type Ca2+ channel is predominantly expressed in the nervous system. To dissect the molecular mechanisms underlying the neuron-specific expression of the P/Q-type channel, we have isolated and characterized the 5'-upstream region of the mouse alpha1A subunit gene. A transcription start site appeared to exist at -269 bp upstream from the start codon as found by 5' RACE analysis. The proximal promoter of the alpha1A subunit gene lacks a typical TATA box, but contains several transcription factor binding sequences, including two Sp1 sites. When linked to a placental alkaline phosphatase (PLAP) reporter gene to examine the promoter activity, the 6.3-kb (-6,273 to +269) 5'-upstream region, but not a smaller 3.0-kb construct (-3, 021 to +269), was able to drive the reporter gene in neuron-like PC12 cells. In contrast, neither of these constructs enhanced the PLAP expression in fibroblast NIH3T3 cells. The sequence between 6.3 and 3.0 kb of the 5'-upstream region did not show promoter activity in either of the cell lines, but enhanced TK promoter activity in PC12 cells, though not in NIH3T3 cells. These results suggest that neuron-specific elements of the alpha1A subunit gene are likely to be located in the distal upstream regions (-6,273 to -3,021) of the 5'-upstream sequence.

3T3 Cells↗

PubMed1998

The upstream, direct repeat sequence of Prague A Rous sarcoma virus is deficient in mediating efficient Gag assembly and particle release.

Rous sarcoma virus (RSV) contains two approximately 135-nt imperfect direct repeats composed of smaller repeats, dr1 (approximately 100 nt) and dr2 (approximately 36 nt), that are between the env and src genes and downstream of src in the 3' untranslated region, respectively. It has previously been shown that a Prague A RSV mutant in which both dr1 sequences are deleted is defective at several points in the virus life cycle, including unspliced RNA and env mRNA stability, unspliced RNA transport, and virus particle assembly. A defect in unspliced RNA transport occurs because a cytoplasmic transport element is present within the dr1. We have suggested that the defect of particle production may arise from the failure of the unspliced RNA to be targeted to sites in the cytoplasm where its translation is favorable for Gag protein assembly. In this report, we have further investigated the function of the direct repeats by comparing virus mutants containing either a single upstream or downstream dr1 sequence. Both mutants were delayed in replication compared to the wild-type; the mutant with a single upstream dr1 (delta DDR) is significantly more defective than the mutant with a single downstream dr1 (delta UDR). While both mutants appear capable of efficiently transporting unspliced RNA to the cytoplasm, the delta DDR mutant with only the upstream dr1 is defective in its ability to support Gag assembly and particle release. The replication defect cannot be repaired by placing the upstream dr1 at the location of the downstream dr1 in the 3' untranslated region. A single point mutation in the upstream dr1 (U to C) restored replication and particle production to near normal levels. The results suggest that unspliced RNA transport and Gag assembly functions may be mediated by different elements within the dr1 and that the Prague A upstream dr1 is defective in the latter but not the former function.

Animals↗

PubMed1989

Three distinct regulatory elements comprise the upstream promoter region of the nopaline synthase gene.

Fine deletion mutants were generated in the upstream control region of the nopaline synthase (nos) promoter to define the position and role of upstream regulatory elements. The results indicated that the 8 bp sequence (CAGAAACC) at -106/-113 and its inverted repeat (GGTTTCTG) at -140/-147 are important for promoter function. The downstream element appears more important than the upstream element since deletion of the former reduced promoter activity more significantly than deletion of the latter. Deletion of the element alone, however, did not abolish promoter function, whereas, deletion of the 10 bp potential Z-DNA-forming (Z) element located between the repeat elements nullified promoter activity. Therefore, it appears that the Z element is an essential upstream regulator and the repeated elements are upstream modulators of the nos promoter. These elements are functionally distinct since alteration of stereospecificity or insertion of short oligonucleotides between the elements did not significantly influence promoter activity. These regulatory elements were unable to function from 200 bp upstream of the CCAAT-TATA box region.

Amino Acid Oxidoreductases↗