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At least 415 records · Page 23Linked to original sources
PubMed2003

Vitamin D growth inhibition of breast cancer cells: gene expression patterns assessed by cDNA microarray.

1,25-Dihydroxyvitamin D3 [1,25(OH)2D3], the active metabolite of vitamin D, is a potent inhibitor of breast cancer cell growth. Although it is evident that 1,25(OH)2D3 inhibits growth of both estrogen receptor alpha-positive [ER alpha(+)] and -negative [ER alpha(-)] breast cancer cells, the cellular pathways contributing to these effects remain unclear. We studied the gene expression patterns in ER alpha(+) MCF-7 and ER alpha(-) MDA MB 231 human breast cancer cells following 1,25(OH)2D3 treatment, using cDNA expression arrays. Both cell lines showed a significant induction of the 1,25(OH)2D3-dependent 24-hydroxylase gene, a marker for the actions of 1,25(OH)2D3. In MCF-7 cells, 51 genes were up-regulated and 19 genes were down-regulated. The up-regulated genes encoded cell adhesion molecules, growth factors/modulators, steroid receptors/co-activators, cytokines, kinases and transcription factors. Of the up-regulated genes, 40% were implicated in cell cycle regulation and apoptosis and included cyclin G1 and cyclin I, p21-activated kinase-1 (PAK-1), p53, retinoblastoma like-2 [Rb2 (p130)], insulin-like growth factor binding protein-5 (IGFBP5) and caspases. Among the down-regulated genes were ER alpha, growth factors, cytokines and several kinases. Some of these results were confirmed by real-time PCR. In MDA MB 231 cells, 20 genes were up-regulated and 13 genes were down-regulated. Very few genes directly implicated in cell cycle regulation were up-regulated. The matrix metalloproteinases formed a major class of genes that were down-regulated in the MDA MB 231 cells. Seven genes were commonly up-regulated in both cell lines and these included transforming growth factor (TGFbeta2) and Rb2 (p130). In conclusion, the gene expression profiles of the two cell lines studied were different with a few overlapping genes suggesting that different cellular pathways might be regulated by 1,25(OH)2D3 to exert its growth inhibitory effects in ER alpha(+) and ER alpha(-) cells.

Antineoplastic Agents↗
PubMed1999

Organization of DNA replication origins in the fission yeast genome.

Eukaryotic DNA replication initiates at multiple points along the chromosomes known as replication origins (ORIs). We have developed a strategy to identify ORIs directly from replication intermediates in the fission yeast Schizosaccharomyces pombe. Mapping of a selection of the novel ORIs onto the genome reveals their preferential localization at intergenic regions upstream from genes. These results are supported by the observation that a large proportion of regions overlapping gene promoters contain active ORIs. Mapping of the genomic ars1 replication origin at nucleotide resolution shows that replication initiates at a defined position immediately upstream from the hus5(+) promoter. Deletion analysis indicates that the regulatory elements required to initiate transcription and replication lie in close proximity, suggesting a possible relationship between both processes in vivo.

Base Sequence↗
PubMed1986

Complete nucleotide sequence of the Escherichia coli ptr gene encoding protease III.

The nucleotide sequence of a 3120 bp region of the E. coli chromosome that includes the entire ptr gene has been determined. The proposed coding region for Protease III is 2889 nucleotides long, which would encode a protein consisting of 962 amino acids with a calculated molecular mass of 107,719 daltons. The predicted primary structure of the protein includes a 23-residue signal sequence, cleavage of which would give rise to a mature protein of molecular mass 105,124 daltons. At its 3' end, the ptr gene overlaps the start of the recB coding sequence by 8 bases, suggesting that these genes may form part of an operon.

Amino Acid Sequence↗
PubMed2004

KpnBI is the prototype of a new family (IE) of bacterial type I restriction-modification system.

KpnBI is a restriction-modification (R-M) system recognized in the GM236 strain of Klebsiella pneumoniae. Here, the KpnBI modification genes were cloned into a plasmid using a modification expression screening method. The modification genes that consist of both hsdM (2631 bp) and hsdS (1344 bp) genes were identified on an 8.2 kb EcoRI chromosomal fragment. These two genes overlap by one base and share the same promoter located upstream of the hsdM gene. Using recently developed plasmid R-M tests and a computer program RM Search, the DNA recognition sequence for the KpnBI enzymes was identified as a new 8 nt sequence containing one degenerate base with a 6 nt spacer, CAAANNNNNNRTCA. From Dam methylation and HindIII sensitivity tests, the methylation loci were predicted to be the italicized third adenine in the 5' specific region and the adenine opposite the italicized thymine in the 3' specific region. Combined with previous sequence data for hsdR, we concluded that the KpnBI system is a typical type I R-M system. The deduced amino acid sequences of the three subunits of the KpnBI system show only limited homologies (25 to 33% identity) at best, to the four previously categorized type I families (IA, IB, IC, and ID). Furthermore, their identity scores to other uncharacterized putative genome type I sequences were 53% at maximum. Therefore, we propose that KpnBI is the prototype of a new 'type IE' family.

Amino Acid Sequence↗
PubMed1984

Construction of a promoter-probe vector for a Bacillus subtilis host by using the trpD+ gene of Bacillus amyloliquefaciens.

The trp gene cluster of Bacillus amyloliquefaciens was found to be structurally similar to that of the Enterobacteriaceae. The translation termination codon of the putative trpE gene and the initiation codon for the putative trpD gene overlap at the trpE-trpD junction, and a promoter for the putative trpC gene is suggested to exist. A promoter-probe vector of Bacillus subtilis, pFTB281, was constructed with a DNA fragment of B. amyloliquefaciens, complementing the trpC and trpD mutations of B. subtilis, a 42-base-pair DNA fragment of M13mp7, and the larger EcoRI-PvuII fragment of pUB110, which confers an autonomous replication function and the kanamycin-resistance phenotype to the chimeric plasmid. pFTB281 has BamHI, EcoRI, and SalI cloning sites in the 5'-upstream portion of the protein-coding region of the putative trpD gene, and the insertion of a certain DNA fragment at any of these sites allowed the plasmid to transform a trpD mutant of B. subtilis to the TrpD+ phenotype. DNA fragments showing the promoter function for the trpD gene were obtained from B. amyloliquefaciens and Saccharomyces cerevisiae chromosomes and rho 11 and lambda phage DNAs, but rarely from the DNAs of Escherichia coli and pBR322.

Amino Acid Sequence↗
PubMed2004

Mimetics of caloric restriction include agonists of lipid-activated nuclear receptors.

The obesity epidemic in industrialized countries is associated with increases in cardiovascular disease (CVD) and certain types of cancer. In animal models, caloric restriction (CR) suppresses these diseases as well as chemical-induced tissue damage. These beneficial effects of CR overlap with those altered by agonists of nuclear receptors (NR) under control of the fasting-responsive transcriptional co-activator, peroxisome proliferator-activated co-activator 1alpha (PGC-1alpha). In a screen for compounds that mimic CR effects in the liver, we found statistically significant overlaps between the CR transcript profile in wild-type mice and the profiles altered by agonists of lipid-activated NR, including peroxisome proliferator-activated receptor alpha (PPARalpha), liver X receptor, and their obligate heterodimer partner, retinoid X receptor. The overlapping genes included those involved in CVD (lipid metabolism and inflammation) and cancer (cell fate). Based on this overlap, we hypothesized that some effects of CR are mediated by PPARalpha. As determined by transcript profiling, 19% of all gene expression changes in wild-type mice were dependent on PPARalpha, including Cyp4a10 and Cyp4a14, involved in fatty acid omega-oxidation, acute phase response genes, and epidermal growth factor receptor but not increases in PGC-1alpha. CR protected the livers of wild-type mice from damage induced by thioacetamide, a liver toxicant and hepatocarcinogen. CR protection was lost in PPARalpha-null mice due to inadequate tissue repair. These results demonstrate that PPARalpha mediates some of the effects of CR and indicate that a pharmacological approach to mimicking many of the beneficial effects of CR may be possible.

Animals↗
PubMedDate not supplied

[Effectiveness of translation coupling in hybrid operons].

The possibility of creating artificial overlappons was studied on the model of two genes, that coding for the N-terminal part of lambda cro protein and the cat of E. coli. To test the dependence of translational coupling efficiency on the intercistronic region a series of recombinant DNA molecules carrying different hybrid operons with partially overlapping genes was constructed. The translational efficiency of the distal to the promoter gene was shown to depend on the intercistronic region structure: overlapping of the AUG codon with the terminating one of the proximal gene in the UGAUG manner is optimal for the translational coupling, and the displacement of AUG at several nucleotides in both directions decreases the translational reinitiation efficiency for the ribosomes, that have synthesised the first gene product.

Acetyltransferases↗
PubMed1997

Iron-regulated haemolysin gene from Edwardsiella tarda.

We have cloned and sequenced the haemolysin gene locus from Edwardsiella tarda (ETH). This region encoded two open reading frames, designated ethA and ethB. ethA is the haemolysin gene consisting of 4782bp encoding a product of 165.3 kDa and ethB is an activation/secretion protein gene of 1677bp that encodes a product of 61.9 kDa. There were two putative ferric uptake regulator (Fur) binding sites on the 5' upstream region of the ethB gene overlapping the promoter region and ribosome-binding site. The haemolysin produced by the cloned gene was secreted by Escherichia coli. The deduced amino acid sequences of the ethA and ethB genes were found to be homologous to those of the haemolysin and activation/secretion proteins of Haemophilus ducreyi, Proteus mirabilis, and Serratia marcescens. E. coli carrying the ethA gene but not the ethB gene completely lost haemolytic activity, although the ethA gene was transcribed. The protein expressed by E. coli carrying a recombinant plasmid which encoded the ethA gene had haemagglutination activity. The EthB protein was necessary for activation of EthA protein (haemolysin). The ethA and ethB genes were very prevalent in haemolytic E. tarda strains isolated from diseased fish. Transcription of the ethB gene was regulated by iron. The ethA and ethB genes were transcribed independently.

Amino Acid Sequence↗
PubMed2000

The transfer region of IncI1 plasmid R64: similarities between R64 tra and legionella icm/dot genes.

The entire nucleotide sequence of the transfer region of IncI1 plasmid R64 was determined together with previously reported sequences. Twenty-two transfer genes, traE-Y and nuc, were newly identified in the present study. The protein products of 17 genes were detected by maxicell experiments or by the T7 RNA polymerase expression system. Mutagenesis experiments indicated that 16 genes were indispensable for R64 transfer both in liquid and on surfaces. In summary, the R64 transfer region located within an approximately 54 kb DNA segment was shown to encode the most complex transfer system so far studied. It contains at least 49 genes and may produce 58 different proteins as a result of shufflon DNA rearrangement and overlapping genes. Among the 49 genes, 23 tra, trb and nik genes have been shown to be indispensable for R64 conjugal transfer in liquid and on surfaces. Twelve additional pil genes are required only for liquid matings. The amino acid sequences of 10 R64 tra/trb products share similarity with those of the icm/dot products of Legionella pneumophila that are responsible for its virulence, suggesting that the R64 transfer and L. pneumophila icm/dot systems have evolved from a common ancestral genetic system.

Amino Acid Sequence↗
PubMed1992

Conservation of the organization of five tightly clustered genes over 600 million years of divergent evolution.

The organization of the mouse surfeit locus is unusual in that it contains six housekeeping genes (Surf-1-Surf-6), which are unrelated by sequence homology, in the tightest mammalian gene cluster thus far described. A maximum of only 73 base pairs separates any two of the four well-characterized genes, and two of the genes overlap at their 3' ends. The direction of transcription of each of the five surfeit genes, Surf-1-Surf-5, alternates with respect to that of its neighbor, suggesting cis-interaction or coregulation between the genes by mechanisms such as the sharing of regulatory elements and/or antisense regulation. The Surf-3 gene has been identified as encoding the ribosomal protein L7a (Rpl7a). We have used the high conservation of the Rpl7a gene to clone the chicken gene and surrounding genomic DNA. The tight clustering and juxtaposition of at least five of the surfeit genes (Surf-1-Surf-5) and their associated CpG-rich islands have been found to be conserved over the 600 million years of divergent evolution that separates birds and mammals. This strongly suggests that the surfeit locus represents a different form of gene cluster in which gene organization may play both a positive and negative regulatory role in gene expression possibly via cis-interactions between the closely spaced genes.

Amino Acid Sequence↗
PubMed2005

Structure and function of the type 3 deiodinase gene.

Thyroid hormones (TH) are essential for normal growth and development in vertebrates, and are important for the maintenance of normal metabolic activity in most tissues of the body. Because the actions of TH result from the binding of 3,3',5'-triiodothyronine (T(3)) to specific nuclear receptors in the target cell, the extent of TH action in a given cell is dependent in part on the intracellular concentration of T(3). The type 3 deiodinase (D3) is a selenoenzyme that inactivates TH by catalyzing their conversion to biologically inactive metabolites. The findings that D3 activity is very high in the pregnant uterus and fetoplacental unit, and that D3-deficient mice exhibit deficits in growth, viability, and fertility strongly suggest that D3 plays an important role in development. The D3 gene (Dio3) is preferentially expressed from the paternally inherited allele and is associated with an overlapping gene transcribed from the opposite DNA strand (Dio3os). D3 mRNA expression and D3 activity are regulated by a number of hormones and growth factors as well as by genomic imprinting. Although some genomic structures appear to mediate some of these effects, many details concerning the function of the Dio3 gene are unresolved. These include the full characterization of the Dio3 and Dio3os genes, the elucidation of the mechanisms responsible for the developmental and tissue-specific patterns observed in Dio3 allelic expression, and the response of the genes to hormones and growth factors. Knowledge of these details will be important for understanding the physiologic function of an enzyme that appears to be critical for normal mammalian development.

Alleles↗
PubMed1985

Structural features of the hisT operon of Escherichia coli K-12.

The DNA sequence of a 2,3-kilobase segment of the E. coli hisT operon was determined. Analysis of the sequence indicated that the upstream gene in the operon encodes a 36,364-dalton polypeptide, which runs aberrantly on SDS-polyacrylamide gels. The distal hisT gene encodes the tRNA modification enzyme, pseudouridine synthase I, which was shown to have a polypeptide molecular mass of 30,399 daltons. The DNA sequence was consistent with the phenotypes and hisT expression of mutant operons. Analysis of the sequence and genetic complementation experiments demonstrated that the upstream and hisT genes are evolutionarily, structurally, and functionally unrelated; however, translation signals for the two genes overlap, which is consistent with genetic evidence suggesting translational coupling. Codon usage in the upstream gene is radically different from the hisT gene and may underlie the differential expression observed from the operon. Gene-inactivation experiments and S1-mapping of in vivo transcripts indicated that the operon contains an additional upstream gene. S1-mapping experiments also confirmed the presence of an internal promoter, which might be stringently controlled. Taken together, these results show that the structure of the hisT operon is complex and suggest that the operon might be regulated at several levels.

Amino Acid Sequence↗
PubMed1989

An additional promoter within the protein-coding region of the psbD-psbC gene cluster in tobacco chloroplast DNA.

Transcription of the psbD-psbC gene cluster in tobacco chloroplasts has been studied. This cluster contains in linear sequence the overlapping genes encoding the D2 and 43 kDa proteins of Photosystem II (psbD and psbC, respectively), and ORF62. Eight major transcripts ranging from 1.5 to 4.4 kb were detected by northern blot analysis. S1 mapping experiments revealed that these multiple transcripts comprise five distinct 5' ends whose precise positions were further determined by primer extension analysis. Two of the five 5' ends were determined to be the transcriptional initiation sites by in vitro capping assays: the main site is located 905 bp upstream from the ATG codon of psbD and the additional site is 194 bp upstream from the first ATG codon of psbC. The latter site and the preceding prokaryotic promoter motif are within the protein-coding region of psbD. The 3' ends of transcripts were determined by S1 mapping.

Base Sequence↗
PubMed1996

RNA editing in the acceptor stem of squid mitochondrial tRNA(Tyr).

In squid (Loligo bleekeri) mitochondria, the two 3'-terminal nucleotides (G72-G73) of the tRNA(Tyr) gene overlap with the two 5'-terminal nucleotides (G1-G2) of the downstream tRNA(Cys) gene. To elucidate the processing mechanism(s) of the tRNA molecules derived from this region, tRNAs were analyzed by sequencing cDNAs synthesized from circularized tRNAs. Nucleotides G1-G2 in tRNA(Cys) appeared to be without post-transcriptional conversion, whereas CCA was post-transcriptionally added to the 3'-terminus. In contrast, in the majority of tRNAs(Tyr), G72-G73 were found to be converted to A72-A73, accompanied by the CCA addition. These results indicate that a precursor of tRNA(Tyr) is processed at U71 and two adenosines are attached prior to the CCA addition. Thus, we suggest that 5' processing of the precursor tRNA dominates 3' processing and maturation of the tRNA is mediated by a polyadenylylation enzyme in the mitochondria, a scenario which is consistent with the editing process proposed in land snail mitochondria. We also obtained intermediates, such as a premature tRNA lacking CCA that terminated at U71 and one with a single adenosine attached at position 72, which support the suggested maturation process. However, although we failed to detect a tRNA(Cys) lacking G1-G2 at the 5'-terminus, we obtained cDNAs for tRNA(Tyr) with G72-G73 and the CCA terminus. This inconsistent result suggests the co-existence of another process(es) in the maturation of these tRNA molecules in squid mitochondria.

Animals↗
PubMed1994

Molecular cloning and sequence analysis of the proBA operon from an extremely thermophilic eubacterium Thermus thermophilus.

A 3.6 kb DNA fragment carrying the Thermus thermophilus proBA region, which encodes the first two steps in the proline biosynthetic pathway, was cloned from the Thermus thermophilus gene library, and its complete nucleotide sequence was determined. The deduced amino acid sequence of gamma-glutamyl kinase (40,657 Da), the product of proB gene, and gamma-glutamyl phosphate reductase (48,747 Da), the product of proA gene, showed 44.1% and 44.4% identity to those of Escherichia coli, respectively. The termination codon of the proB gene and the initiation codon of the proA gene overlapped by 2 bp. A possible transcriptional termination structure was found downstream of the proA gene but not downstream of the proB gene. These results indicate that the proBA genes of T. thermophilus form a single operon as in E. coli.

Aldehyde Oxidoreductases↗
PubMed1990

Sequence and genetic organization of a Zymomonas mobilis gene cluster that encodes several enzymes of glucose metabolism.

The Zymomonas mobilis genes that encode glucose-6-phosphate dehydrogenase (zwf), 6-phosphogluconate dehydratase (edd), and glucokinase (glk) were cloned independently by genetic complementation of specific defects in Escherichia coli metabolism. The identity of these cloned genes was confirmed by various biochemical means. Nucleotide sequence analysis established that these three genes are clustered on the genome and revealed an additional open reading frame in this region that has significant amino acid identity to the E. coli xylose-proton symporter and the human glucose transporter. On the basis of this evidence and structural analysis of the deduced primary amino acid sequence, this gene is believed to encode the Z. mobilis glucose-facilitated diffusion protein, glf. The four genes in the 6-kb cluster are organized in the order glf, zwf, edd, glk. The glf and zwf genes are separated by 146 bp. The zwf and edd genes overlap by 8 bp, and their expression may be translationally coupled. The edd and glk genes are separated by 203 bp. The glk gene is followed by tandem transcriptional terminators. The four genes appear to be organized in an operon. Such an arrangement of the genes that govern glucose uptake and the first three steps of the Entner-Doudoroff glycolytic pathway provides the organism with a mechanism for carefully regulating the levels of the enzymes that control carbon flux into the pathway.

Amino Acid Sequence↗
PubMed1996

Genomic cloning and characterization of the human thrombin receptor gene. Structural similarity to the proteinase activated receptor-2 gene.

The seven-transmembrane segment thrombin receptor (TR) represents the prototype of a putative family of proteolytically cleaved receptors that may include the proteinase activated receptor-2. A panel of somatic cell hybrids retaining distinct portions of human chromosome 5 were used to establish that the human TR gene is present as a single-copy locus within the region 5q11.2 -->q13.3, confirming our previous localization using fluorescent in situ hybridization analysis. To further characterize the TR gene, overlapping clones from a human genomic library were isolated. Genomic analysis confirmed that the TR gene is of limited complexity, spanning approximately 27 kilobases and containing two exons separated by a large approximately 22-kilobase intron. The larger second exon contains the majority of the coding sequence and the thrombin cleavage site, remarkably similar to the organization of the proteinase activated receptor-2 gene in which the putative cleavage site is also contained within the large second exon. Primer extension analysis using two 30-mer oligonucleotide primers known to be contained within the first exon identified the predominant transcription initiation site 351 base pairs upstream from the initiator methionine in both human umbilical vein endothelial and human erythroleukemia cells. Sequence analysis of the 5'-flanking region revealed the TR promoter to be TATA-less, although nucleic acid motifspotentially involved in transcriptional gene regulation were evident and include a GATA motif, octamer enhancer sequences, AP-2-like sites, and Sp1 sites. These data provide evidence for remarkable similarity at the gene level between both proteolytically cleaved receptors described to date.

Base Sequence↗
PubMed1994

Disruption of virus movement confers broad-spectrum resistance against systemic infection by plant viruses with a triple gene block.

White clover mosaic virus strain O (WClMV-O), species of the Potexvirus genus, contains a set of three partially overlapping genes (the triple gene block) that encodes nonvirion proteins of 26 kDa, 13 kDa, and 7 kDa. These proteins are necessary for cell-to-cell movement in plants but not for replication. The WClMV-O 13-kDa gene was mutated (to 13*) in a region of the gene that is conserved in all viruses known to possess triple-gene-block proteins. All 10 13* transgenic lines of Nicotiana benthamiana designed to express the mutated movement protein were shown to be resistant to systemic infection by WClMV-O at 1 microgram of WClMV virions per ml, whereas all plants from susceptible control lines became systemically infected. Of the 13* transgenic lines, 3 selected for their abundant seed supply were shown to be resistant to systemic infection when challenged by inoculation with three different WClMV strains (O, M, and J) or with WClMV-O RNA at 10 micrograms/ml. Most plants were also resistant to systemic infection at inoculum concentrations up to 250 micrograms of WClMV virions per ml. In addition, the three 13* transgenic plant lines were found to be resistant to systemic infection with two other members of the Potexvirus group, potato virus X and narcissus mosaic virus, and the Carlavirus potato virus S but not to be resistant to tobacco mosaic virus of the Tobamovirus group. These results indicate that virus resistance can be engineered into transgenic plants by expression of dominant negative mutant forms of triple-gene-block movement proteins.

Base Sequence↗