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At least 271 records · Page 15Linked to original sources
PubMed1989

Identification of enhancer elements in the upstream region of the nuclear photosynthetic gene ST-LS1.

The nuclear gene ST-LS1 from potato encodes a 10-kilodalton protein that is a component of the oxygen-evolving complex of photosystem II. Analysis of the expression of a reporter gene driven by chimeric promoters, consisting of ST-LS1 upstream sequences and a truncated cauliflower mosaic virus 35S promoter, suggests that a strong positive regulatory element is located between position -345 and -261, whereas both the region -261 to +11 and the more upstream region -1600 to -530 are devoid of autonomous strong positive elements detectable by this approach. The ST-LS1 upstream region contains redundant elements conferring light-regulated and organ-specific expression, one of them being located between position -130 and +11. In addition, enhancer-like sequences conferring light-regulated as well as organ-specific expression to heterologous promoters were identified. These sequences are functional not only when located 5'-upstream of the coding region but also when placed 3'-downstream of the polyadenylation signal, thus representing one of the first examples of a plant gene-derived enhancer being able to induce a truncated heterologous promoter from a position 3'-downstream of the transcription unit.

Cell Nucleus↗
PubMed1989

Multiple domains exist within the upstream activator sequence of the octopine synthase gene.

It is known that a 16-base pair palindrome (ACGTAAGCGCTTACGT) located upstream of the ocs gene can activate a maize adh1 promoter in a transient expression system [Ellis et al. (1987). EMBO J. 6, 11-16; Ellis et al. (1987). EMBO J. 6, 3203-3208]. We have determined that this palindrome is also essential for ocs promoter activity in tobacco calli. In addition, sequences immediately adjacent to this palindrome, both 5' and 3', modulate its activity. The palindrome is sensitive to the differentiated state of the plant cells in which it resides; it is active in calli and the leaves of small shoots but is inactive in the leaves of rooted plants. We have tested upstream sequences from two other T-DNA genes that have homology to this palindrome for their ability to activate the octopine synthase promoter in tobacco calli. The upstream region from the mannopine synthase gene can activate the octopine synthase promoter, but an upstream region from the gene implicated in octopine and nopaline secretion cannot activate the promoter.

Agrobacterium tumefaciens↗
PubMed1993

Analysis of the 5'-upstream promoter region of human involucrin gene: activation by 12-O-tetradecanoylphorbol-13-acetate.

Involucrin is one of the precursor proteins of keratinocyte cornified envelope. Although the formation of the cornified envelope is induced by tumor-promoting phorbol esters, the effect of 12-O-tetradecanoylphorbol-13-acetate (TPA) on the involucrin gene expression remains unknown. We have isolated a 5'-upstream region of human involucrin gene and examined its TPA-dependent promoter activity. The involucrin upstream region with the untranslated first exon was connected to chloramphenicol acetyltransferase (CAT)-involucrin promoter expression vector (INV-CAT) and was transfected into fetal rat keratinizing epidermal (FRSK) cells. The INV-CAT-transfected FRSK cells showed considerable CAT activity that was significantly augmented by the treatment of cells with TPA. FRSK cells that were transfected with a reversely oriented 5'-upstream sequence revealed little CAT activity and did not respond to TPA. The effect of TPA was significantly inhibited by the protein kinase C inhibitor 1-(5-isoquinoline-sulfonyl)-2-methyl piperazine dihydrochloride (H-7). Other protein kinase C activators (1-oleoyl-2-acetylglycerol and mezerein) also induced the INV-CAT promoter activity, whereas 4-O-methyl phorbol myristate acetate, a very weak protein kinase C activator, had only a slight effect. Analysis of the nucleotide sequence of the 5'-upstream region detected several 5'-TGANTCAA-3' sequences that are highly conserved TPA-response elements (TRE). Cotransfection of both c-jun and c-fos expression vectors with the INV-CAT vector into FRSK cells resulted in increased CAT activity. Cotransfection of either the c-jun or c-fos vector singly with the INV-CAT vector into FRSK cells had negligible effects. Dexamethasone significantly inhibited the TPA-induced promoter activity in the INV-CAT-transfected FRSK cells. These results indicate that involucrin gene expression is positively controlled by TPA through the activation of the protein kinase C/TRE system.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1993

The function of the upstream region of the sigma 54-dependent Klebsiella pneumoniae nifL promoter is sensitive to DNA supercoiling.

The positive control protein NTRC activates transcription from the sigma 54-dependent nifL and glnAp2 promoters of Klebsiella pneumoniae by binding to upstream enhancer-like sequences and contacting downstream bound sigma 54-RNA polymerase via looping of the intervening DNA. In contrast to the glnAp2 promoter, the activity of the nifL promoter is very sensitive to changes in DNA supercoiling both in vivo and in vitro. We have shown previously that the downstream elements of the nifL promoter are involved in the supercoiling response. In this study we find that the upstream region of nifL influences the supercoiling response of a hybrid nifL-glnAp2 promoter both in vivo and in vitro, demonstrating that the nifL upstream region also confers supercoiling sensitivity. DNA supercoiling did not appear to influence binding of NTRC to its sites in the nifL upstream region, suggesting that another function of this region, most probably DNA loop formation, is sensitive to changes in DNA topology.

Bacterial Proteins↗
PubMed1995

Effects of upstream deletions on light- and oxygen-regulated bacterio-opsin gene expression in Halobacterium halobium.

The bacterio-opsin gene (bop) of Halobacterium halobium is located within a cluster with three other genes. Growth conditions of high light intensity and low oxygen tension induce bop gene cluster expression. To identify putative regulatory factor binding sites upstream of the bop gene, we have compared sequences upstream of the bop gene with the corresponding sequences from two other genes in the bop gene cluster. Conserved sequence motifs were observed which may mediate the effect of high light intensity and/or low oxygen tension on bop gene expression. Based on these motifs, a set of mutants was constructed which contained deletions upstream of the bop gene. These constructs were tested in a host strain where bop gene expression is independent of oxygen regulation and in another strain where it is regulated by oxygen and light. The minimal upstream sequence required for both light- and oxygen-regulated bop gene expression was determined to be 54 bp.

Bacteriorhodopsins↗
PubMed1996

Functional in vivo studies of the Neurospora crassa cys-14 gene upstream region: importance of CYS3-binding sites for regulated expression.

Sulphate transport in Neurospora crassa is achieved by two distinct sulphate permeases, I and II, encoded by the cys-13 and cys-14 genes, respectively. The synthesis of both sulphate permeases is subject to sulphur repression and requires the global positive-acting regulatory protein CYS3, CYS3, a bZIP DNA binding protein, regulates cys-14 expression at the transcriptional level and binds in vitro specifically to three DNA-recognition sites, A, B, and C, in the cys-14 upstream region. In vivo functional analysis of the cys-14 promoter was carried out with 5' deletions and by deletions or mutations of CYS3 DNA-binding sites. The most distal CYS3-binding site, C, located 1.4kb upstream of the transcriptional start site, is necessary and sufficient to mediate strong transcriptional activation by CYS3; moreover, site C was able to function equally well when it was located at variable distances upstream of the cys-14 gene. Site B, located 1 kb upstream, alone is able to support a moderate degree of cys-14 expression. Site A is not required and does not appear to play any functional role in cys-14 expression, even though it is in close proximity to the transcriptional start site. The presence of multiple copies of CYS3-binding elements A or B in the cys-14 promoter results in a parallel increase of regulated gene expression. When a transforming cys-14 gene becomes integrated at ectopic locations in the host genome, it can be expressed in an unregulated fashion, presumably by coming under the control of other promoter elements. Our results also suggested that at least one enzyme in the sulphate catabolic pathway requires a functional CYS3 protein for expression.

Anion Transport Proteins↗
PubMed1988

Sequence analysis of murine cytokeratin endo A (no. 8) cDNA. Evidence for mRNA species initiated upstream of the normal 5' end in PCC4 cells.

Keratin 8 (Endo A) is expressed in simple epithelia, together with keratin 18 (Endo B). Filaments formed by this keratin pair are the first cytoskeletal elements induced during mouse embryogenesis. We have isolated Endo A cDNA clones from lambda gt11 libraries prepared with mRNA isolated from PCC4 embryonal carcinoma (EC) cells. Sequencing of three overlapping cDNAs and of a genomic clone allowed us to determine the complete sequence of the Endo A message. Analysis of the protein sequence deduced showed that the Endo A protein presented all the characteristics of intermediate filaments, including an alpha-helical central rod domain and nonhelical N- and C-termini. In the rod domain, the degree of similarity to the other members of the basic keratin family was high. A high degree of homology to keratin 8 of other species was observed, even in the non-helical domains. During these analyses, we found clones extending upstream of the normal 5' end of the mRNA. Sequence comparison between these cDNAs and the 5' upstream region of the Endo A gene suggested that they corresponded to transcripts initiated at an upstream alternative promoter. These observations supported previous results showing the presence of Endo A transcripts initiated upstream of the normal 5' end in mouse morulae and blastocysts.

Amino Acid Sequence↗
PubMed1990

Interaction of upstream stimulatory factor with the human heme oxygenase gene promoter.

Upstream stimulatory factor (USF), originally identified in HeLa cells, interacts with the upstream promoter sequence of adenovirus 2 major late promoter (Ad2MLP) and activates its transcription. USF is present in uninfected HeLa cells and appears to be involved in the transcription of cellular genes related to stress. Recently, we have proposed that the rat heme oxygenase gene, newly identified heat-shock protein gene, is regulated at least in partly by a rat homolog of USF [Sato, M., Fukushi, Y., Ishizawa, S., Okinaga, S., Muller, R.M. & Shibahara, S. (1989) J. Biol. Chem. 264, 10251-10260]. We therefore confirm that the heme oxygenase gene is expressed in HeLa cells and its expression is increased by cadmium, suggesting that human heme oxygenase is a stress protein similar to the metallothioneins. Using partially purified USF from HeLa cells, we show that USF binds to the human heme oxygenase gene promoter and stimulates its cell-free transcription. The cis-acting element, identified as CACGTGACCCG, is located 34 bp upstream from the transcription initiation site, and contains the core sequence of the upstream promoter sequence of Ad2MLP. We propose that USF contributes to the transcription of the human heme oxygenase gene.

Adenoviridae↗
PubMed2004

Polymorphisms in the 5'-upstream region of the PKCbeta gene in Japanese patients with Type 2 diabetes.

AIMS: Protein kinase C (PKC), a serine/threonine kinase, is known to be activated in various tissues under hyperglycaemic conditions. Notably, PKCbeta, a member of the conventional PKC group, is the predominant isoform detected in vascular tissues and could be involved in the development of diabetic vascular complications. In the present study, we investigated genetic variations in the 5'-upstream region of the PKCbeta gene to assess their possible relation to vascular complications in diabetic patients. METHODS: Variations upstream from the PKCbeta gene (-1066/+256) were examined in 60 Type 2 diabetic patients using a cycle sequencing method. Screening of detected variations was performed in 204 Type 2 diabetic patients and 160 healthy controls. RESULTS: Five single nucleotide polymorphisms; C(-238)G, C(-287)T, A(-348)G, C(-546)G, and C(-853)T, were identified in the upstream region. The C(-287)T and A(-348)G polymorphisms were in perfect linkage disequilibrium. There were no significant differences in genotype or allele frequencies of the five polymorphisms among the diabetic patients and healthy subjects. However, both -238GG and -287CC (-348GG) homozygotes showed significantly higher frequencies of macrovascular disease compared with patients with other genotypes. Further, an electrophoretic mobility shift assay revealed that the -238G fragment had a five-fold higher affinity for transcription factor Sp1 when compared with -238C. CONCLUSIONS: The C(-238)G and C(-287)T-A(-348)G polymorphisms in the 5'-upstream region of the PKCbeta gene may have an effect on the susceptibility of diabetic vascular complications through an alteration of tissue PKCbeta density or function.

Base Sequence↗
PubMed2006

Vicissitudes of benefit sharing of crop genetic resources: downstream and upstream.

In this article, we will first give a historic overview of the concept of benefit sharing and its appearance in official agreements, particularly with respect to crop genetic resources. It will become clear that, at present, benefit sharing is primarily considered as an instrument of compensation or exchange, and thus refers to commutative justice. However, we believe that such a narrow interpretation of benefit sharing disregards, and even undermines, much of its (historical) content and potency, especially where crop genetic resources are concerned. We argue that benefit sharing should not be based merely on commutative justice but rather on a broader model that is also grounded in the concept of distributive justice. This has repercussions for the application of benefit sharing, which we try to clarify by distinguishing between downstream and upstream benefit sharing. Upstream benefit sharing is not so much inspired by compensation for actions done, or the distribution downstream of benefits developed, but by the idea of shared decision-making on the research and development of resources fundamental to human welfare. Going upstream in the research process of crop genetic resources, and determining research agendas and improving crops according to the needs of the poor, benefit sharing may well be a tool to contribute to world food security and global justice. We concretize our ideas on upstream benefit sharing by introducing a set of criteria that determine the success of consultations on agricultural research agenda setting.

Biodiversity↗
PubMedDate not supplied

Conserved expression domains for genes upstream and within the HoxA and HoxD clusters suggests a long-range enhancer existed before cluster duplication.

The posterior HoxA and HoxD genes are essential in appendicular development. Studies have demonstrated that a "distal limb enhancer," remotely located upstream of the HoxD complex, is required to drive embryonic autopod expression of the posterior Hox genes as well as the two additional non-Hox genes in the region: Evx2 and Lnp. Our work demonstrates a similar mode of regulation for Hoxa13 and four upstream genes: Evx1, Hibadh, Tax1bp, and Jaz1. These genes all show embryonic (E11.5-E13.5) distal limb and genital bud expression, suggesting the existence of a nearby enhancer influencing the expression of a domain of genes. Comparative sequence analysis between homologous human and mouse genomic sequence upstream of Hoxa13 revealed a remote 2.25-kb conserved noncoding sequence (mmA13CNS) within the fourth intron of the Hibadh gene. mmA13CNS shares a common 131-bp core identity within a conserved noncoding sequence upstream of Hoxd13, which is located within the previously identified distal limb enhancer critical region. To test the function of this conserved sequence, we created mmA13CNS-Hsp86-lacZ transgenic mice. mmA13CNS directed a wide range of tissue expression, including the central nervous system, developing olfactory tissue, limb, and genital bud. Limb and genital bud expression directed by mmA13CNS is not identical to the patterns exhibited by Hoxa13/Evx1/Hibadh/Tax1bp1/Jaz1, suggesting that mmA13CNS is not sufficient to fully recapitulate their expression in those tissues. The Evx1- and Evx2-like central nervous system expression observed in these mice suggests that the long-range regulatory element(s) for the Hox cluster existed before the cluster duplication.

Animals↗
PubMed2001

Novel group of virulence activators within the AraC family that are not restricted to upstream binding sites.

Several regulators within the AraC family control the expression of genes known or thought to be required for virulence of bacterial pathogens. One of these, Rns, activates transcription from an unprecedented variety of binding-site locations. Although nearly all prokaryotic activators bind within a small region upstream and adjacent to the promoter that they regulate, Rns does not bind within this region to activate its own promoter, Prns. Instead, to activate Prns, Rns requires one binding site 224.5 bp upstream and one downstream of the transcription start site. We show in this study that several other AraC family activators recognize the same binding sites as Rns and share with it the ability to utilize a downstream binding site. Like Rns, other members of this group of activators positively regulate the expression of virulence factors in pathogenic bacteria. These regulators are also able to activate transcription from promoter-proximal upstream binding sites since they are able to substitute for Rns at Pcoo, a promoter with only upstream binding sites. Thus, Rns is the prototype for a group of regulators, which include CfaR, VirF, AggR, and CsvR and which activate transcription from locations that are more diverse than those of any other known activator.

Amino Acid Sequence↗
PubMed1986

Expression of Escherichia coli infC: identification of a promoter in an upstream thrS coding sequence.

infC, the gene which codes for translation initiation factor 3, is situated in a cluster in the genome of Escherichia coli with genes for several other components of the translation apparatus. Only three nucleotides separate the termination codon of thrS from the initiation codon of infC. This implies that infC is either cotranscribed with thrS from a thrS promoter or that the transcriptional signals for infC are embedded within the upstream thrS coding region. In the present work, several plasmids have been constructed which encompass infC and various amounts of the upstream thrS sequence. The ability of the plasmid DNA, or derived restriction fragments, to direct the synthesis of initiation factor 3 was tested in an in vitro DNA-dependent coupled transcription-translation system and in plasmid-transformed maxicells. The results indicate that initiation factor 3 is synthesized in the absence of the thrS promoter. A promoter whose presence is sufficient for the expression of infC has been localized to an 89-base-pair region which lies 178 to 267 base pairs upstream of the infC initiation codon. S1 nuclease mapping of in vivo transcripts confirms that a transcription initiation site is located in this region. These studies demonstrate that infC can be transcribed from a promoter within the upstream thrS coding sequence.

Amino Acyl-tRNA Synthetases↗
PubMed1988

Characterization of the upstream region of the formate dehydrogenase operon of Methanobacterium formicicum.

The fdhA and fdhB genes of Methanobacterium formicicum, which code for the alpha and beta subunits of formate dehydrogenase, were cotranscribed as part of a large transcript. By using Northern (RNA) gel blot analysis, the transcription start site was located within a 1.6-kilobase BglII-NcoI fragment 4.3 kilobases upstream from the fdhA gene. The precise transcription start site within the fragment was determined with the aid of primer extension analysis and S1 nuclease protection studies. A putative promoter sequence for structural genes of methanogenic archaebacteria is proposed based on a comparison of DNA sequences of the upstream region of methanogen operons for which transcription initiation sites are known. Comparison of the DNA sequence of the upstream region of the fdh operon of M. formicicum with the sequence upstream of the fdhF gene of Escherichia coli revealed regions of considerable identity.

Aldehyde Oxidoreductases↗
PubMed1989

Transcription from the P1 promoters of Micromonospora echinospora in the absence of native upstream DNA sequences.

We demonstrated previously that the 0.4-kilobase DNA fragment from Micromonospora echinospora contains multiple tandem promoters, P1a, P1b, P1c, and P2, which are also functional when cloned into Streptomyces lividans. We now show by in vitro transcription with Streptomyces RNA polymerase that each of these promoters is an authentic initiation site, rather than a processing site for transcripts which initiate further upstream. The DNA sequence requirements for the closely spaced promoters P1a, P1b, and P1c, which are coordinately induced during stationary phase in M. echinospora, were examined by deletional analysis in S. lividans. The P1a and P1b promoters were functional despite deletion of native sequences 5 and 17 base pairs upstream of each initiation site, respectively. Thus, P1a and P1b had greatly reduced upstream DNA sequence requirements compared with typical procaryotic promoters. In contrast, transcription from promoter P1c was significantly decreased when native sequences 34 base pairs upstream were replaced.

Base Sequence↗
PubMed1990

A sequence-specific DNA-binding factor (VF1) from Anabaena sp. strain PCC 7120 vegetative cells binds to three adjacent sites in the xisA upstream region.

A DNA-binding factor (VF1) partially purified from Anabaena sp. strain PCC 7120 vegetative cell extracts by heparin-Sepharose chromatography was found to have affinity for the xisA upstream region. The xisA gene is required for excision of an 11-kilobase element from the nifD gene during heterocyst differentiation. Previous studies of the xisA upstream sequences demonstrated that deletion of this region is required for the expression of xisA from heterologous promoters in vegetative cells. Mobility shift assays with a labeled 250-base-pair fragment containing the binding sites revealed three distinct DNA-protein complexes. Competition experiments showed that VF1 also bound to the upstream sequences of the rbcL and glnA genes, but the rbcL and glnA fragments showed only single complexes in mobility shift assays. The upstream region of the nifH gene formed a weak complex with VF1. DNase footprinting and deletion analysis of the xisA binding site mapped the binding to a 66-base-pair region containing three repeats of the consensus recognition sequence ACATT.

Base Sequence↗
PubMed1993

Detection of an A-factor-responsive protein that binds to the upstream activation sequence of strR, a regulatory gene for streptomycin biosynthesis in Streptomyces griseus.

DNA-binding assays using mobility shift polyacrylamide gel electrophoresis revealed the presence of a protein that specifically bound to a restriction fragment -288 to -191 bp upstream from the transcriptional start point of strR, a regulatory gene for streptomycin biosynthesis in Streptomyces griseus. The binding site corresponded to an upstream activation sequence predicted from the results of in vivo promoter assays. The binding was greatly enhanced by 5 mM Mg2+. This binding was detected with the protein source only from the wild-type strain and not from an A-factor-deficient mutant strain. The exogenous supplementation of A-factor to the A-factor-deficient mutant strain caused the appearance of the protein in the DNA-binding assay. A synthetic nucleotide 52 bp in length (region from -293 to -242), which was synthesized on the basis of data obtained from both retardation assays with dissected DNA fragments and in vivo promoter assays, was retarded by the A-factor-dependent protein. In addition to this A-factor-dependent protein, at least three proteins with different recognition site affinities capable of binding to the upstream region of the strR promoter were detected. The binding of one of these proteins to both sides of the upstream activation sequence bound by the A-factor-dependent protein was completely abolished in the presence of ATP and Mg2+ in the incubation mixture. The region bound by these proteins showed anomalous electrophoretic mobility, like that of a bent DNA molecule, which is probably caused by the presence of many blocks consisting of A and T. The region bound by these proteins was found to be transcribed in the orientation opposite to that of strR.

4-Butyrolactone↗
PubMed1994

Positive and negative regulation of sequences upstream of the form II cbb CO2 fixation operon of Rhodobacter sphaeroides.

The unlinked form I and form II Calvin cycle CO2 fixation (cbb) operons of the photosynthetic bacterium Rhodobacter sphaeroides are located on different genetic elements, yet both operons are positively regulated by the transcription activator protein CbbR, the product of the cbbR gene located immediately upstream of the form I operon. By employing deletion mutagenesis, and a newly constructed promoter probe vector, the form II operon promoter (cbbFIIp) and three other promoters (Up, Vp, and Wp) were localized within 2.1 kb upstream of the form II operon. Mutations in both cbbR and the first gene of the form I operon (cbbFI) elicited both positive and negative responses when transcriptional fusions controlled by these four promoters were examined. With the exception of Wp, all these upstream promoters were repressed by oxygen. In addition, these promoters were associated with open reading frames of unknown function whose deduced amino acid sequences showed no significant relationship to proteins in current databases. The results of these experiments suggest that the promoter sequences and genes upstream of the form II cbb operon may be intimately involved with control of the cbb regulon of this photosynthetic organism.

Amino Acid Sequence↗