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Showing papers on "Transcription (biology) published in 1982"


Journal ArticleDOI
TL;DR: A series of recombinant genomes which directed expression of the enzyme chloramphenicol acetyltransferase (CAT) in mammalian cells provided a uniquely convenient system for monitoring the expression of foreign DNAs in tissue culture cells.
Abstract: We constructed a series of recombinant genomes which directed expression of the enzyme chloramphenicol acetyltransferase (CAT) in mammalian cells. The prototype recombinant in this series, pSV2-cat, consisted of the beta-lactamase gene and origin of replication from pBR322 coupled to a simian virus 40 (SV40) early transcription region into which CAT coding sequences were inserted. Readily measured levels of CAT accumulated within 48 h after the introduction of pSV2-cat DNA into African green monkey kidney CV-1 cells. Because endogenous CAT activity is not present in CV-1 or other mammalian cells, and because rapid, sensitive assays for CAT activity are available, these recombinants provided a uniquely convenient system for monitoring the expression of foreign DNAs in tissue culture cells. To demonstrate the usefulness of this system, we constructed derivatives of pSV2-cat from which part or all of the SV40 promoter region was removed. Deletion of one copy of the 72-base-pair repeat sequence in the SV40 promoter caused no significant decrease in CAT synthesis in monkey kidney CV-1 cells; however, an additional deletion of 50 base pairs from the second copy of the repeats reduced CAT synthesis to 11% of its level in the wild type. We also constructed a recombinant, pSV0-cat, in which the entire SV40 promoter region was removed and a unique HindIII site was substituted for the insertion of other promoter sequences.

7,438 citations


Journal ArticleDOI
23 Jul 1982-Science
TL;DR: Transcriptional control signals of a model eukaryotic protein-coding gene have been identified by a new procedure of in vitro mutagenesis that allows small clusters of nucleotide residues to be substituted in a site-directed manner without causing the addition or deletion of other sequences.
Abstract: Transcriptional control signals of a model eukaryotic protein-coding gene have been identified by a new procedure of in vitro mutagenesis. This method allows small clusters of nucleotide residues to be substituted in a site-directed manner without causing the addition or deletion of other sequences. Transcription assays of a systematic series of these clustered point mutants have led to the identification of three distinct control signals located within the 105-nucleotide residues immediately upstream from the point where transcription begins.

1,131 citations


Journal ArticleDOI
01 Oct 1982-Cell
TL;DR: It is shown here that a plasmid containing a cloned gene coding for a yeast leucine tRNA comes under developmental control when injected into cleaving eggs, suggesting that the MBT is triggered by the DNA through titration of suppressor components present in the egg.

1,105 citations


Journal ArticleDOI
01 Sep 1982-Cell
TL;DR: It seems that the upstream element of the hsp 70 promoter is analogous to that of other promoters, but is only functional in heat-shocked cells.

897 citations


Journal ArticleDOI
01 Mar 1982-Cell
TL;DR: A consensus sequence is uncovered between the region deleted in cyc1-512 and the 3' nontranslated regions of some but not all yeast genes, and the possible role of this sequence in transcription termination is discussed.

858 citations


Journal ArticleDOI
TL;DR: The nucleotide sequence of twoBacillus subtilis promoters (veg andtms) that are utilized by the principal form of B. subtilIS RNA polymerase found in vegetative cells is determined and sequences are compared to those of several previously reported Bacillus promoters.
Abstract: We have determined the nucleotide sequence of twoBacillus subtilis promoters (veg andtms) that are utilized by the principal form ofB subtilis RNA polymerase found in vegetative cells (σ55-RNA polymerase) and have compared our sequences to those of several previously reportedBacillus promoters Hexanucleotide sequences centered approximately 35 (the “-35” region) and 10 (the “-10” region) base pairs upstream from theveg andtms transcription startpoints (and separated by 17 base pairs) corresponded closely to the consensus hexanucleotides (TTGACA and TATAAT) attributed toEscherichia coli promoters Conformity to the preferred -35 and -10 sequences may not be sufficient to promote efficient utilization byB subtilis RNA polymerase, however, since three promoters (veg, tms andE coli tac) that conform to these sequences and that are utilized efficiently byE coli RNA polymerase were used with highly varied efficiencies byB subtilis RNA polymerase

816 citations


Journal ArticleDOI
TL;DR: Experiments employing strains inducible (GAL80) or constitutive (gal80) for GAL10 expression indicate that an additional component of glucose repression is inducer exclusion.
Abstract: We have identified the promoter region of the GAL10 gene (whose product is UDP-galactose epimerase) of Saccharomyces cerevisiae; this promoter mediates galactose induction of transcription in conjunction with the product of the GAL4 regulatory gene. This identification was achieved by excising a 365-base-pair fragment of GAL10 leader DNA with a GAL10 proximal endpoint greater than 100 base pairs upstream of the transcriptional start site and substituting it in place of the upstream activation site of the CYC1 (iso-1-cytochrome c) promoter [Guarente, L. & Ptashne, M. (1981) Proc. Natl. Acad. Sci. USA 78, 2199-2203]. The hybrid promoter is composed of DNA encoding CYC1 mRNA start sites and the GAL segment upstream of these sites. This promoter is regulated in a manner analogous to GAL10; i.e., it is induced by galactose and responds to mutations in the GAL4 and GAL80 regulatory loci. The activity of the hybrid promoter requires sequences in the region of the CYC1 mRNA start sites but does not require a precise spacing between these sequences and the GAL segment. The transposed GAL segment appears not to contain sequences that mediate glucose repression. Thus, the picture of the GAL10 promoter that emerges is one of an upstream activation site that responds to the GAL4 product plus galactose, and a region of transcription initiation that may contain sequences that mediate glucose repression. Experiments employing strains inducible (GAL80) or constitutive (gal80) for GAL10 expression indicate that an additional component of glucose repression is inducer exclusion.

506 citations


Journal ArticleDOI
TL;DR: There appeared to be a correlation between levels of c-myb expression and hemopoietic activity of the tissues and cells examined, which suggests that c- myb may be expressed primarily in immature hemopOietic cells.
Abstract: The oncogenes (v-onc genes) of rapidly transforming retroviruses have homologs (c-onc genes) in the genomes of normal cells. In this study, we characterized and quantitated transcription from four c-onc genes, c-myb, c-myc, c-erb, and c-src, in a variety of chicken cells and tissues. Electrophoretic analysis of polyadenylated RNA, followed by transfer to nitrocellulose and hybridization to cloned onc probes showed that c-myb, c-myc, and c-src each give rise to a single mature transcript, whereas c-erb gives rise to multiple transcripts (B. Vennstrom and J. M. Bishop, Cell, in press) which vary in abundance among different cells and tissues. Transcription from c-myb, c-myc, c-erb, and c-src was quantitated by a "dot-blot" hybridization assay. We found that c-myc, c-erb, and c-src transcription could be detected in nearly all cells and tissues examined, whereas c-myb transcription was detected only in some hemopoietic cells; these cells, however, belong to several different lineages. Thus, in no case was expression of a c-onc gene restricted to a single cell lineage. There appeared to be a correlation between levels of c-myb expression and hemopoietic activity of the tissues and cells examined, which suggests that c-myb may be expressed primarily in immature hemopoietic cells. An examination of c-onc RNA levels in target cells and tissues for viruses carrying the corresponding v-onc genes revealed no obvious correlation, direct or inverse, between susceptibility to transformation by a given v-onc gene and expression of the homologous c-onc gene.

380 citations


Journal ArticleDOI
01 Oct 1982-Cell
TL;DR: Four RNA transcripts encoded by cauliflower mosaic virus DNA have been detected in the polyadenylated RNA from virus-infected turnip leaves, two of which have the same 5' termini, at nucleotide 7435.

347 citations


Journal ArticleDOI
01 May 1982-Cell
TL;DR: Pulse-chase experiments performed both in vivo and in vitro support the idea that most newly synthesized 5S rRNA molecules are transiently associated with the La protein, and suggest that this protein plays an essential role in the synthesis or maturation of all class III transcripts.

322 citations


Journal ArticleDOI
TL;DR: The transcription of the human cytomegalovirus genome was investigated at immediate early, early, and late times after infection and it is proposed that expression of the immediate early viral genes is required to transcribe theEarly viral genes in the long repeat and adjacent sequences.
Abstract: The transcription of the human cytomegalovirus genome was investigated at immediate early, early, and late times after infection. Viral RNAs associated with either the whole cell, the nucleus, the cytoplasm, or the polyribosomes were analyzed. At immediate early times, i.e., in the absence of de novo viral protein synthesis, the viral RNA in high abundance originated from a region of the long unique section of the prototype arrangement of the viral genome (0.660 to 0.770 map units). The viral RNA in low abundance originated from the long repeat sequences (0.010 to 0.035 and 0.795 to 0.825 map units) and a region in the long unique section (0.201 to 0.260 map units). Viral RNAs associated with the polyribosomes as polyadenylated RNA were mapped to these restricted regions of the viral genome and characterized according to size class in kilobases. At 24 h after infection in the presence of an inhibitor of viral DNA replication, i.e., at early times, the stable viral RNAs in highest abundance mapped in the long repeat sequences. Viral RNAs at intermediate abundance under these conditions mapped in two regions of the long unique section of the viral genome (0.325 to 0.460 and 0.685 to 0.770 map units). Stable viral RNAs that were associated with the polyribosomes in high abundance as polyadenylated RNA orginated from the long repeat sequences, but not from the long unique section of the viral genome. An analysis of whole-cell RNA at late times (72 h) indicated that the abundant transcription was in the regions of the long unique sequences (0.325 to 0.460 and 0.660 to 0.685 map units), and transcription of intermediate abundance was from the long repeat sequences. However, stable viral mRNA's derived from the long repeat sequences were associated with the polyribosomes at late times after infection. In addition, mRNA's originating from the long and short unique sequences were found associated with the polyribosomes at higher relative concentration than at early times after infection. It is proposed that expression of the immediate early viral genes is required to transcribe the early viral genes in the long repeat and adjacent sequences. These sequences are also transcribed at late times after infection while viral DNA synthesis continues. The expression of viral genes in most of the long and short unique sequences appears to require viral DNA replication.

Journal ArticleDOI
01 Jul 1982-Cell
TL;DR: Analysis of nuclear RNA suggests that the beta 0 transcript is inefficiently spliced and that the removal of the two intervening sequences is coupled.

Journal ArticleDOI
TL;DR: The exons in c-myc may define two functional domains in the gene and may therefore facilitate the dissection of the different oncogenic potentials of the MC29 virus.
Abstract: The chicken genome contains nucleotide sequences homologous to transforming genes (oncogenes) of a number of avian retroviruses. We have isolated chicken DNA (c-myc) that is homologous to the oncogene (v-myc) of the avian myelocytomatosis virus MC29 and have compared the structures of the cellular and viral genes. Results from restriction endonuclease mapping of c-myc and from analysis of heteroduplexes between the DNAs of the cellular and viral genes show that c-myc is homologous to 1,500 nucleotides in v-myc DNA. This homologous region is interrupted in c-myc by an intron-like sequence of 1,100 nucleotides which is absent from v-myc. Nuclear RNA from normal chicken cells contains at least five species of transcripts from c-myc ranging from 2.5 to 6.5 kilobases in length. By contrast, cytoplasm contains only the 2.5-kilobase c-myc RNA. These features of the c-myc gene and its nuclear transcripts are characteristic of normal cellular genes and suggest that the myc gene is of cellular rather than viral origin. The exons in c-myc may define two functional domains in the gene and may therefore facilitate the dissection of the different oncogenic potentials of the MC29 virus.

Journal ArticleDOI
TL;DR: The precise site of initiation of transcription of alpha gene 0 within the inverted b sequences of the L component of viral DNA is reported on.
Abstract: Previous studies have shown that herpes simplex virus genes form three groups, alpha, beta, and gamma, whose expression is coordinately regulated and sequentially ordered in a cascade fashion. Chimeric genes constructed by fusion of the coding and 5' nontranslated leader sequences of the thymidine kinase (TK) gene to the sequences upstream from the site of initiation of transcription of alpha genes 4 and 27 are regulated as alpha genes and are induced in cells converted to TK+ phenotype by infection with TK- virus. In alpha gene 4 (S. Mackem and B. Roizman, Proc. Natl. Acad. Sci. U.S.A. 79:4917-4921, 1982), both the promoter and the regulatory region are separable and movable. The promoter permits expression but not induction when fused to TK in the noncoding leader region of the gene. The regulator, when fused to the promoter of an expressible but noninducible portion of the natural beta TK, renders the gene inducible as an alpha gene; it consists of multiple regulatory units acting cumulatively. In this paper, we report on the precise site of initiation of transcription of alpha gene 0 within the inverted b sequences of the L component of viral DNA. We also report the following. (i) The chimeric gene consisting of the coding and 5' nontranslated leader regions of the TK gene fused to portions of the domain of alpha gene 0 extending largely upstream from the site of initiation of transcription of alpha gene 0 was regulated in the same fashion as the alpha 4- and alpha 27-TK chimeras. The regulatory region in the alpha gene 0 is largely upstream from nucleotide - 140. (ii) The promoter-regulatory regions of alpha genes 0, 4, and 27 share TATA sequences, A + T-rich (consensus) sequences occurring in regulating regions of alpha genes 0 and 4 in more than one copy, and multiple G + C-rich inverted repeats. The relation of these sequences to the function of the promoter-regulatory regions of the alpha genes is discussed.

Journal ArticleDOI
TL;DR: The presence of two initiation sites for H-strand transcription can be correlated with other types of evidence that point to two different transcription events leading to the synthesis of a polycistronic molecule corresponding to the almost entire H strand and to the synthesisation of the rRNA species.
Abstract: The initiation sites for heavy (H) and light (L) strand transcription in HeLa cell mitochondrial DNA have been investigated by mapping experiments utilizing in vitro "capped" mitochondrial RNA molecules or nascent RNA chains. Mitochondrial poly(A)-containing RNA molecules were labeled at their 5' ends with [alpha-32P]GTP and guanylyltransferase ("capping" enzyme) and mapped on the mitochondrial genome by DNA transfer hybridization and S1 nuclease protection experiments. A mapping site for the capped 5' ends was found on the H strand very near to the 5' terminus of the 12S rRNA gene, and another site was found on the L strand very near to the 5' terminus of the 7S RNA coding sequence. In parallel experiments, the 5' ends of the nascent chains isolated from mitochondrial DNA transcription complexes were similarly mapped very near to the 5' termini of the 12S rRNA gene and of the 7S RNA coding sequence. The in vitro capped RNA molecules and the nascent chains thus presumably identify the same transcriptional initiation sites on the H strand and the L strand. The occurrence of a second possible initiation site for H-strand transcription 90-110 nucleotides upstream of that described above--i.e., 20-40 nucleotides upstream of the tRNAPhe gene--had been previously indicated by a mapping analysis of the nascent RNA chains and has been confirmed in the present work. The presence of two initiation sites for H-strand transcription can be correlated with other types of evidence that point to two different transcription events leading to the synthesis of a polycistronic molecule corresponding to the almost entire H strand and to the synthesis of the rRNA species.

Patent
24 Nov 1982
TL;DR: A cloning vector comprises a replication system from a bovine papilloma virus, a regulatory system including the promoter and terminator from a human metallothionein II gene, and a prokaryotic replication system as discussed by the authors.
Abstract: A cloning vector comprises a replication system from a bovine papilloma virus, a regulatory system including the promoter and terminator from a human metallothionein II gene, and a prokaryotic replication system. The vector is useful for transforming mammalian cells where it is maintained as an automonously replicating episome. Transcription may be regulated by the presence of heavy metals and/or glucocorticoid hormones in the growth media.

Journal ArticleDOI
TL;DR: Findings suggest that estradiol increases prolactin mRNA levels by increasing the transcription of the prolact in pituitary cells, and not on the other hand as previously suggested.

Journal ArticleDOI
TL;DR: SP6 RNA polymerase possesses a stringent promoter specificity similar to, but distinct from that of the other phage RNA polymerases, and is also highly active in synthesis of poly(rG) with poly(dI) .

Journal ArticleDOI
01 Jul 1982-Cell
TL;DR: The possibility that promoter occlusion at two lambda promoters, Pint and PR', might play a role in the sequential expression of viral functions is discussed.

Journal ArticleDOI
01 Feb 1982-Cell
TL;DR: It is proposed that stable transcription complexes may play an important role in the maintenance of the differentiated state in eucaryotic cells.

Journal ArticleDOI
TL;DR: High resolution analysis of runoff transcripts showed that the reconstituted system initiated transcription precisely at the adenovirus major late and early region IV promoters.

Journal ArticleDOI
TL;DR: The copy number of plasmids containing the ColE1 replicon is affected by changes in the transcriptional activity within the plasmid if these changes lead to transcriptional readthrough into the replication region towards the promoter priming DNA replication.
Abstract: The copy number of plasmids containing the ColE1 replicon is affected by changes in the transcriptional activity within the plasmid if these changes lead to transcriptional readthrough into the replication region towards the promoter priming DNA replication. Such readthrough e.g., from the tet region in pBR322 not only causes overproduction of a peptide known to affect the copy number negatively but also appears to interfere negatively with the replication of the plasmid itself. The proper placement of efficient transcriptional terminators prevents such interference and permits the stable integration of strong promoters. Due to this termination effect, up to 9-fold differences in plasmid copy number were observed, depending upon the particular growth conditions. The higher copy number is of course reflected by higher yields of plasmid-specified gene products indicating the relevance of the above effects for studies of gene expression.

Journal ArticleDOI
TL;DR: The results suggest that the stimulation of accumulation of small subunit and light‐harvesting complex mRNAs by exposure of Pisum seedlings to light is mediated by an increase in transcription rather than by a decrease in RNA degradation.
Abstract: Nuclei isolated from both light-grown and dark-grown leaves of Pisum sativum by Percoll density gradient centrifugation incorporate labelled UTP into RNA when supplemented with the other three nucleoside triphosphates. The RNA is heterodisperse, with transcripts up to at least 25S in size. Among these transcripts are sequences hybridizing to cloned DNA probes for wheat rRNA and two abundant chloroplast polypeptides of Pisum, viz. the small subunit of ribulose bisphosphate carboxylase and a polypeptide of the light-harvesting chlorophyll a/b binding complex. Transcription of small subunit and light-harvesting complex sequences is greater (18-fold and 9-fold, respectively) in nuclei from light-grown leaves than in nuclei from dark-grown leaves. Transcription of ribosomal genes, by contrast, is only doubled by growth in the light. Small subunit and light-harvesting complex sequences transcribed in dark-grown nuclei are not degraded in a 120 min chase. These results suggest that the stimulation of accumulation of small subunit and light-harvesting complex mRNAs by exposure of Pisum seedlings to light is mediated by an increase in transcription rather than by a decrease in RNA degradation.

Journal ArticleDOI
14 Jan 1982-Nature
TL;DR: Comparison of the level of β-globin transcripts in a variety of deletion mutants shows that for efficient transcription, both the ATA or Goldberg–Hogness box, and a region between 100 and 58 base pairs in front of the site at which transcription is initiated, are required.
Abstract: The DNA sequences required for the expression of the rabbit-beta-globin gene in vivo have been examined. A variety of mutant rabbit beta-globin gene templates were linked to a simian virus 40-plasmid recombinant and introduced into HeLa cells; in these conditions the rabbit beta-globin gene is expressed from its own promoter. Comparison of the level of beta-globin transcripts in a variety of deletion mutants shows that for efficient transcription, both the ATA or Goldberg-Hogness box, and a region between 100 and 58 base pairs in front of the site at which transcription is initiated, are required. Deletion of either of these regions results in a decrease in the level of beta-globin transcripts by an order of magnitude; deletion of the ATA box causes an additional loss in the specificity of the site of initiation of RNA synthesis. The DNA sequences downstream from the ATA box, including the natural beta-globin mRNA cap site, are dispensable for transcription in vivo.

Journal ArticleDOI
01 Dec 1982-Cell
TL;DR: The almost exclusive location of strong contact points on the noncoding strand of the gene suggests how transcription events could proceed repeatedly along a gene that is assembled into a stable transcription complex.

Journal ArticleDOI
TL;DR: The results confirmed that the most abundant cytoplasmic RNA species in such cells is specified by a small region of the genome defined by the EcoRI J fragment.
Abstract: Cytoplasmic RNA prepared from five lymphoid cell lines and a Burkitt lymphoma biopsy was radioactively labeled in vitro and hybridized to cloned EcoRI restriction endonuclease fragments of B95-8 Epstein-Barr virus DNA. The results confirmed that the most abundant cytoplasmic RNA species in such cells is specified by a small region of the genome defined by the EcoRI J fragment. Detailed mapping experiments precisely localized these transcripts within the sequence of the rightmost one-third of the EcoRI J fragment. DNA sequencing suggested that this region of the Epstein-Barr virus genome is unable to code for protein. The major early transcripts consisted of two non-polyadenylated RNA species, each about 170 nucleotides in length. They were both transcribed off the same strand of the DNA and showed significant sequence homology with each other. The coding sequences of the two small RNAs contained potential intragenic control regions for RNA polymerase III.

Journal ArticleDOI
TL;DR: Introduction of a segment containing the SV40 early polyadenylation signal into the 3' end of the DH FR cDNA gene generated a recombinant capable of transforming cells to the DHFR+ phenotype with at least a 10-fold increase in efficiency, demonstrating the necessity for an efficient polyadenyation signal.
Abstract: Dihydrofolate reductase (DHFR) modular genes have been constructed with segments containing the adenovirus major late promoter, a 3' splice site from a variable region immunoglobulin gene, a DHFR cDNA, and portions of the simian virus 40 (SV40) genome. DNA-mediated transfer of these genes transformed Chinese hamster ovary DHFR- cells to the DHFR+ phenotype. Transformants contained one to several copies of the transfected DNA integrated into the host genome. Clones subjected to growth in increasing concentrations of methotrexate eventually gave rise to lines containing several hundred copies of the transforming DNA. Analysis of the DHFR mRNA produced in amplified lines indicated the following. (i) All clones utilize the adenovirus major late promoter for transcription initiation. (ii) A hybrid intron formed by the 5' splice site of the adenovirus major late leader and a 3' splice site from a variable-region immunoglobulin gene is properly excised. (iii) The mRNA is not efficiently polyadenylated at sequences in the 3' end of the DHFR cDNA but rather uses polyadenylation signals downstream from the DHFR cDNA. Three independent clones produce a DHFR mRNA containing SV40 or pBR322 and SV40 sequences, and the RNA is polyadenylated at the SV40 late polyadenylation site. Another clone has recombined into cellular DNA and apparently uses a cellular sequence for polyadenylation. Introduction of a segment containing the SV40 early polyadenylation signal into the 3' end of the DHFR cDNA gene generated a recombinant capable of transforming cells to the DHFR+ phenotype with at least a 10-fold increase in efficiency, demonstrating the necessity for an efficient polyadenylation signal. Attachment of a DNA segment containing the transcription enhancer (72-base pair repeat) of SV40 further increased the biological activity of the modular DHFR gene 50- to 100-fold.

Journal ArticleDOI
TL;DR: It is proposed that a polypeptide of 63 amino acids whose sequence is completely conserved in pMBl and ColE1 is the rop gene product and that it regulates plasmid DNA replication by modulating the initiation of transcription of the primer RNA precursor.
Abstract: A 600-base-pair region essential for ColE1 and pMBl plasmid replication contains two promoters responsible for the synthesis of two RNA molecules central to copy number control. One promoter directs synthesis of the primer RNA precursor. The second promoter directs the synthesis of a small RNA molecule, RNAl, which acts in trans to inhibit processing of the RNA primer precursor. We have fused each promoter to the beta-galactosidase structural gene contained in a lambda phage. Expression of the RNAl promoter in lysogens is not influenced by the presence of wild-type pMBl or ColEl plasmids residing in the cell. Transcription from the RNA primer promoter, however, is repressed by the product of a trans-acting plasmid gene product, which we have designated rop (for repressor of primer). The rop gene maps downstream from the replication origin in a region that encodes a polypeptide of 63 amino acids whose sequence is completely conserved in pMBl and ColE1. We propose that this polypeptide is the rop gene product and that it regulates plasmid DNA replication by modulating the initiation of transcription of the primer RNA precursor.

Journal Article
TL;DR: The results show that the cloned mouse metallothionein-I gene retains its cadmium inducibility and that the regulation of this gene occurs primarily at the transcriptional level.
Abstract: Cadmium and other heavy metals induce metallothionein synthesis in mammalian cells. We have cloned a mouse metallothionein-I gene from a cadmium-resistant line of Ltk- cells and inserted it into viral and plasmid SV40 vectors. These recombinant molecules were introduced into cultured monkey cells and the expression of the foreign metallothionein gene was analyzed, at the RNA level, in the presence and absence of cadmium. The results show that the cloned gene retains its cadmium inducibility and that the regulation of this gene occurs primarily at the transcriptional level.

Journal ArticleDOI
11 Mar 1982-Nature
TL;DR: Data presented here show that in vitro transcription of ribosomal DNA isolated from mouse, human and a protozoan requires completely homologous components, and none of the three active cell-free systems is capable of correct initiation on the nonhomologous templates.
Abstract: Eukaryotic cells possess three distinct nuclear DNA-dependent RNA polymerases which are responsible for transcription of different sets of genes (for review see refs 1, 2). Recently, cell-free transcription systems have been developed which faithfully initiate transcription of isolated genes by the corresponding RNA polymerase in the presence of crude cellular extracts. These cellular extracts supply additional components required for specific transcription3–6. Successful in vitro systems for transcription of RNA polymerase II or III genes were developed using either heterologous or homologous components7–11. In contrast, an analogous cell-free system for the RNA polymerase I transcription unit from mouse has been shown to be active only with homologous extracts from mouse cells6. Data presented here show that in vitro transcription of ribosomal DNA isolated from mouse, human and a protozoan requires completely homologous components. None of the three active cell-free systems is capable of correct initiation on the nonhomologous templates. Further, supplementation of mouse extracts with purified protozoan RNA polymerase I failed to result in specific transcription of the protozoan rDNA, suggesting that the species specificity of pre-ribosomal RNA synthesis resides, in part, in the transcription factors.