Showing papers on "Gene expression published in 1974"
TL;DR: The data discussed herein demonstrate the great variation in target-tissue response that can occur after administration of steroid hormones, and direct quantitative evidence that sex steroids cause a net increase in the intracellular amounts of specific mRNA molecules in target tissues is provided.
Abstract: The data discussed herein demonstrate the great variation in target-tissue response that can occur after administration of steroid hormones. The female sex steroids can exert regulatory effects on the synthesis, activity, and possibly even the degradation of tissue enzymes and structural proteins. Each response, nevertheless, appears to be dependent on the synthesis of nuclear RNA. In many instances, the steroid actually promotes a qualitative change in the base composition and sequence of the RNA synthesized by the target cell, implying a specific effect on gene transcription. Most important is our direct quantitative evidence that sex steroids cause a net increase in the intracellular amounts of specific mRNA molecules in target tissues. It thus appears that we are discovering a pattern of steroid hormone action which includes (Fig. 1): (i) uptake of the hormone by the target cell and binding to a specific cytoplasmic receptor protein; (ii) transport of the steroid-receptor complex to the nucleus; (iii) binding of this "active" complex to specific "acceptor" sites on the genome (chromatin DNA and acidic protein); (iv) activation of the transcriptional apparatus resulting in the appearance of new RNA species which includes specific mRNA's; (v) transport of the hormone-induced RNA to the cytoplasm resulting in synthesis of new proteins on cytoplasmic ribosomes; and (vi) the occurrence of the specific steroid-mediated "functional response" characteristic of that particular target tissue. To elucidate fully the mechanism of steroid hormone action we must study the biochemistry of the process by which information held by the steroid hormone-receptor complex is transferred to the nuclear transcription apparatus. If our assumptions are correct, we should ultimately be able to discover how this hormone-receptor complex exerts a specific regulatory effect on nuclear RNA metabolism. Such regulation might be achieved (i) by direct effects on chromatin template leading to increased gene transcription and thus RNA synthesis; (ii) by activation of the polymerase complex itself; (iii) by inhibition of RNA breakdown; or (iv) by intranuclear processing of large precursor molecules so that smaller biologically active sequences are produced, and (v) by transport of RNA from the nucleus to the cytoplasmic sites of cellular protein synthesis.
866 citations
TL;DR: It is apparent that, in addition to regulating the transcription of defined genome loci, the nonhistone chromosomal proteins include enzymes that have a general function, proteins that are involved in determining the structure of chromatin, as well as proteins that serve as recognition sites for binding of regulatory macromolecules.
Abstract: Evidence from several model systems suggests that nonhistone chromosomal proteins may regulate gene expression in eukaryotic cells. The data indicate that the synthesis of new species of nonhistone chromosomal proteins as well as modifications of preexisting nonhistone chromosomal proteins are involved in the control of transcription. However, from the vast number of proteins included in this class, it is apparent that, in addition to regulating the transcription of defined genome loci, the nonhistone chromosomal proteins include enzymes that have a general function, proteins that are involved in determining the structure of chromatin, as well as proteins that serve as recognition sites for binding of regulatory macromolecules. The presence of a nucleoplasmic pool of nonhistone chromosomal proteins which may exchange with the chromatin has also been reported (89). While it is clear that the nonhistone chromosomal proteins play a key role in the regulation of gene expression, the exact manner in which they interact with the genome to initiate, modify, or augment the transcription of specific RNA molecules remains to be resolved.
415 citations
166 citations
TL;DR: Cytoplasmic RNA extracted from human tissue culture cells infected with adenovirus type 2 was used to program protein synthesis in a cell-free system derived from mammalian cells, demonstrating seven size classes of RNA each of which programmed the synthesis of only one or two virus-specific polypeptides.
Abstract: Cytoplasmic RNA extracted from human tissue culture cells infected with adenovirus type 2 was used to program protein synthesis in a cell-free system derived from mammalian cells. Analysis of the protein product by polyacrylamide gel electrophoresis revealed ten adenovirus-specific polypeptides. Five of these were further identified by analysis of tryptic peptides. Translation of RNA fractionated by sedimentation through sucrose gradients containing formamide demonstrated seven size classes of RNA, each of which programmed the synthesis of only one or two virus-specific polypeptides. Six of the virus-specific polypeptides were translated from RNAs much larger than expected for the size of the polypeptide.
134 citations
TL;DR: The transcripts of native and reconstituted chromatins were indistinguishable in respect of both the total yield of the RNA and the fractional yield of globin-specific sequences.
Abstract: Chromatin was reconstituted from the purified DNA and histones of chicken erythrocytes and the nonhistone proteins of either chicken reticulocytes or chicken liver. Reconstituted chromatins, native chicken reticulocyte chromatin, and free DNA were transcribed with Escherichia coli RNA polymerase and the concentrations of globin-specific sequences in the RNA products were measured by hybridization with [3H]DNA complementary to chicken globin messenger RNA. Reticulocyte, but not liver, nonhistone proteins were shown to activate the globin genes in reconstituted erythrocyte chromatin. The transcripts of native and reconstituted chromatins were indistinguishable in respect of both the total yield of the RNA and the fractional yield of globin-specific sequences.
126 citations
TL;DR: Analysis of human competitor proteins with homologous assay systems of viral core proteins and corresponding antisera showed that all, including the normal tissue extracts, appear similar to core proteins of known viruses, especially the RD 114 and woolly monkey species.
Abstract: Partially purified fractions of human tissues have been analyzed by competition radioimmunoassay for the presence of two of the principle structural components of type-C RNA viruses, the major core protein (p27 to p30) and the major envelope glycopeptides (gp69/71). Screening of tissues was carried out by use of a heterologous assay system of 125 I-labeled Rauscher murine virus p30 antigen and anti-RD 114 virus serum which was found to detect a class of interspecies determinants common to murine, feline, and primate viruses. A competitor with the same apparent affinity for antibody binding as that of purified viral core proteins was found in relatively high concentration in tissues from patients with systemic lupus erythematosus, in some neoplastic tissues, and also in normal human tissues. This competitor from a lupus spleen chromatographed on phosphocellulose and showed size fractionation during gel filtration similar to known p27 to p30 viral proteins. An immunologically reactive protein was also demonstrated by immunodiffusion and by immunoprecipitation of 125 I-labeled human protein with anti-RD 114 p28 serum. Analysis of these human competitor proteins with homologous assay systems of viral core proteins and corresponding antisera showed that all, including the normal tissue extracts, appear similar to core proteins of known viruses, especially the RD 114 and woolly monkey species. A hypothesis suggested by these data is that many, if not all, humans harbor at least part of the genome of one or more type-C viruses, the properties of which are similar to those of viruses from other mammalian species, particularly primates. Images
113 citations
99 citations
TL;DR: The presence of cycloheximide during the early phase of adenovirus 2 replication causes an increase in the virus-specific content of newly synthesized mRNA, which appears to be caused by a reduction in synthesis of cellular cytoplasmic mRNA.
Abstract: The presence of cycloheximide during the early phase of adenovirus 2 replication causes an increase in the virus-specific content of newly synthesized mRNA. The total cytoplasmic RNA from control cultures labeled 2 to 5 h after infection hybridized to viral DNA 0.8%, whereas RNA synthesized in the presence of cycloheximide annealed 6%. Cytosine arabinoside, an inhibitor of DNA synthesis, did not affect the percent hybridization to viral DNA. Oligo(dT)-cellulose chromatography was used to purify the portion of cytoplasmic RNA containing poly(A). The poly(A)-containing RNA from cultures labeled in the presence of cycloheximide hybridized to viral DNA 32% as compared to 2.2% for RNA from control cultures. Hybridization-inhibition experiments between RNAs from control- and cycloheximide-treated cultures demonstrated that the cultures treated with cycloheximide did not have an increased content of viral RNA or a new class of viral RNA sequences. Therefore, the increased hybridization appears to be caused by a reduction in synthesis of cellular cytoplasmic mRNA. Nucleoplasmic RNAs lacking and containing poly(A) were annealed to viral DNA. For both classes, RNA from cultures treated with cycloheximide hybridized 5- to 10-fold more than RNA from control-infected cultures. Therefore, the increased hybridization of cytoplasmic RNA synthesized in the presence of cycloheximide is caused either by reduced transcription of the cellular genome or by greatly increased instability of cellular heterogeneous nuclear RNA.
85 citations
TL;DR: There is control of transcription of the globin gene in this cell and probably also control at a post-transcriptional level, according to complementary DNA synthesised on a template of mouse globin mRNA.
85 citations
TL;DR: The evidence indicates that the membrane-bound poly(A)-containing RNA is a product of the nuclear poly (A)containing RNA and is the precursor of the polysomal poly( A)-containingRNA.
Abstract: Three different classes of poly(A)-containing RNA have been identified in the yeast system. They have been characterized by their kinetics of labeling, by their localization in the nuclear, membranous, and soluble cytoplasmic fractions, and by their size. The evidence indicates that the membrane-bound poly(A)-containing RNA is a product of the nuclear poly(A)-containing RNA and is the precursor of the polysomal poly(A)-containing RNA. In addition, it has been demonstrated that transport of mRNA is coupled with protein synthesis.
TL;DR: It is shown that an inhibitor of translation, which appears to be a protein associated with the ribosomes, accumulates in extracts of interferon treated L cells and can be eliminated by the addition of purified fractions of mammalian transfer RNA.
TL;DR: The viabilities of coliphage T4 mutants demonstrate that neither alteration nor modification is essential for growth in Escherichia coli B/r, and these specific phage-induced structural changes in the host RNA polymerase are not fundamental to the control of gene expression during T4 development.
TL;DR: This report proves that gene 55 codes directly for the 22,000 molecular weight polymerase-binding protein, which requires a full-sized, but not an active, gene 55 product.
TL;DR: Two independent procedures demonstrated that early cytoplasmic viral RNA is composed of two sequence classes, class I which is absent or present in greatly reduced quantities at 18 h, and class II which persists throughout the infection.
Abstract: The RNA sequences and RNA size classes transcribed early in productive infection with adenovirus 2 were analyzed by RNA-DNA hybridization. Two independent procedures demonstrated that early cytoplasmic viral RNA is composed of two sequence classes, class I which is absent or present in greatly reduced quantities at 18 h, and class II which persists throughout the infection. When the sequences in early viral RNA were analyzed by hybridization-inhibition studies, the hybridization of early [3H]RNA was inhibited only 50% by RNA from cultures harvested late (18 h) in infection. Liquid hybridizations with radioactive viral DNA confirmed that early RNA includes two classes. Duplex formation of RNA with 32P-labeled viral DNA was assayed by hydroxylapatite chromatography and resistance to S1 nuclease digestion. Both methods showed that the cytoplasmic RNA present early in infection annealed 12 to 15% of the viral DNA; late cytoplasmic RNA hybridized 21 to 25% of the DNA. Mixtures of early plus late cytoplasmic RNAs hybridized 30 to 34% of the viral DNA, demonstrating the reduced concentration of early class I RNA in the late RNA preparations. Experiments were performed to correlate class I and class II early RNA with size-fractionated cytoplasmic RNA synthesized early in infection. Fractionation of RNA by gel electrophoresis or sucrose gradient centrifugation confirmed three major size classes, 12 to 15S, 19 to 20S, and 26S. Total cytoplasmic RNA and RNA selected on the basis of poly(A) content contained the same size classes of viral RNA. In standard electrophoresis conditions, the 19 to 20S viral RNA could be resolved into two size classes, and the distribution of 12 to 15S RNA also indicated the presence of more than one size component. Hybridization-inhibition studies under nonsaturating conditions were performed with 26S, 19 to 20S, and 12 to 15S viral RNAs fractionated by gel electrophoresis. Late RNA inhibited the hybridization of 26S RNA only 20%, 19 to 20S RNA was inhibited 45%, and 12 to 15S RNA was inhibited 50%. When 18 to 19S and 12 to 15S viral RNAs purified by sucrose gradient centrifugation were similarly analyzed, late RNA inhibited hybridization of 18 to 19S RNA 50%, and the annealing of 12 to 15S RNA was inhibited 70%.
TL;DR: Results show that transcription of phage DNA occurs in vivo well after the occurrence of host changes that inhibit phage replication, suggesting that the changes in the template specificity of host RNA polymerase probably cannot account for the inability of phages to multiply at early stages of sporulation.
TL;DR: A protein kinase from rat liver nuclei stimulates RNA synthesis andosphorylation of RNA polymerase appears to be another mechanism by which protein kinases regulate gene expression.
TL;DR: The change in the interferon sensitivity of SV40 T antigen synthesis suggests that the expression of at least this portion of the SV40 genetic information in Ad2(+)ND(4) is under Ad2 genetic control, which suggests that transcription of the genetic information for SV 40 T antigen is initiated in a region of Ad2 DNA.
Abstract: The effect of interferon on simian virus 40 (SV40) and adenovirus 2 (Ad2) T antigen synthesis has been examined in cells infected with SV40, with Ad2, and with a nondefective Ad2-SV40 hybrid virus, Ad2(+)ND(4). The induction of SV40 T antigen by SV40 was highly sensitive to interferon, whereas the induction of Ad2 T-antigen by Ad2 was resistant. This difference in interferon sensitivity was also noted in cells simultaneously infected with both viruses. However, the induction of SV40 T antigen by Ad2(+)ND(4), which contains covalently linked SV40 and Ad2 DNAs, was as resistant to interferon as the induction of Ad2 T antigen. This change in the interferon sensitivity of SV40 T antigen synthesis suggests that the expression of at least this portion of the SV40 genetic information in Ad2(+)ND(4) is under Ad2 genetic control. When RNA extracted from Ad2(+)ND(4)-infected cells was examined by means of sequential hybridization with Ad2 DNA, elution, and rehybridization with SV40 DNA, 27% of the SV40-specific RNA was found to be linked to Ad2 RNA. No such linkage was detected in control mixtures of Ad2 and SV40 RNAs. The presence of Ad2 and SV40 nucleotide sequences in the same RNA molecule implies that, in Ad2(+)ND(4) infection, transcription is initiated in the DNA of one virus (Ad2 or SV40) and continues without interruption across the point of junction into the DNA of the other virus. Furthermore, the interferon resistance of Ad2(+)ND(4)-induced SV40 T antigen synthesis suggests that transcription of the genetic information for SV40 T antigen is initiated in a region of Ad2 DNA.
TL;DR: It is shown that virus induction by cycloheximide requires de novo RNA synthesis during but not after drug exposure and suggested that the required RNA species may be that of the virus itself, consistent with the hypothesis that translational inhibitors prevent synthesis of a labile protein whose normal action is to inhibit viral RNA transcription or to cause degradation of viral RNA.
Abstract: Mouse cells contain the genetic information for multiple endogenous type-C RNA viruses. The mechanisms by which the cell controls expression of these naturally integrated viruses are not yet known. Recently, chemicals that inhibit protein synthesis have been shown to induce a specific type-C virus at high frequency from BALB/c mouse embryo cells. In the present studies, virus activation in response to a representative translational inhibitor, cycloheximide, is demonstrated to be transient, with virus release primarily occurring within the first 12-24 hr following drug exposure. Analysis of virus-specific RNA in cells by molecular hybridization revealed an absolute increase in viral RNA concentration in cycloheximide-treated cells. This was blocked by simultaneous exposure of the cells to actinomycin D. Further, inhibition of RNA synthesis during but not subsequent to cycloheximide exposure prevented virus activation. These findings show that virus induction by cycloheximide requires de novo RNA synthesis during but not after drug exposure and suggest that the required RNA species may be that of the virus itself. The present results are consistent with the hypothesis that translational inhibitors prevent synthesis of a labile protein whose normal action is to inhibit viral RNA transcription or to cause degradation of viral RNA.
TL;DR: The distribution of RNA labelled in vivo compared with that of RNA polymerase activities suggested that RNA synthesized in vivo is rapidly translocated from its site of synthesis to some other sites within the nucleus.
Journal Article•
TL;DR: The mammalian trophoblast cell, its unique position in the lineages of cells derived from the zygote, the gene sets apparently involved in the production of this specialized cell phenotype, and the evidence that its mitosis and/or phenotypic expression can be regulated by the uterine milieu during normal reproduction are proposed.
Abstract: Summary Although ontogeny may ultimately be best described as a continuous process and although gene repression may never be absolute, there is, nonetheless, considerable experimental and conceptual justification for designating more-or-less discrete phases in ontogeny which can be defined in terms of characteristically predominant sets of expressed genes. The concept of developmental phases is compatible with evidence demonstrating temporal sequences of only partially overlapping transcriptive sets, such as appear to occur during early embryogenesis, as well as with evidence demonstrating a progressive restriction in transcriptional activity within a given set, as appears to occur during terminal differentiation within a cell lineage. The phase-specific gene products that have been identified or inferred in developing systems include, in addition to detectable antigens, RNA polymerases, transfer RNA species, transfer RNA methyltransferases and their inhibitors, aminoacyl transfer RNA synthetases, ribosomal proteins, and “masking” proteins associated with the cell membrane. Since the regulation of gene expression in eukaryotes appears to involve integrated gene sets rather than individual genes, we may predict that these gene products are members of such sets whose other members remain to be identified. We may further predict that the reexpression of these gene products during the malignant transformation of adult cells may not involve a random derepression of individual genes, but rather an orderly switch from one integrated gene set to another, such as occurs in transdetermination. Work in our laboratory has focused on genetic expression during the early phases of mammalian embryogenesis. We have found that, even in multipotential embryonic cells, the transcription of nonrepetitive DNA sequences is actually more restricted than that in several adult organs. During the period of intense organogenesis, transcription of nonrepetitive DNA sequences is more widespread, as might be expected, but it is also evident that genes expressed earlier are repressed at this later stage. There are demonstrable losses of discrete protein synthetic capacities even during the first days of embryogenesis, and marked decreases in transfer RNA methyltransferase activity. Attention is called to the mammalian trophoblast cell, its unique position in the lineages of cells derived from the zygote, the gene sets apparently involved in the production of this specialized cell phenotype, and the evidence that its mitosis and/or phenotypic expression can be, and frequently are, regulated by the uterine milieu during normal reproduction. It is proposed that this natural phenomenon may provide clues toward solving clinical problems involving the multiplication and differentiation of the malignant cell.
TL;DR: The results suggest that the polypeptide composition and the products of the cell-associated RNA polymerase are similar to those of the RNA transcriptase associated with influenza virus particles.
Abstract: The products synthesized in vitro by an RNA-dependent RNA polymerase isolated from influenza virus-infected BHK21-F cells were analyzed by velocity sedimentation, annealing techniques, and acrylamide-agarose gel electrophoresis. Approximately 50% of the RNA synthesized in vitro remains associated with the 50 to 70S ribonucleoprotein complex containing polymerase activity; the remainder of the RNA polymerase product sediments heterogeneously with a peak at 13S. At least 90% of the in vitro product hybridizes with virion RNA. If polypeptides are labeled early in the growth cycle, both the P and NP polypeptides are detected in the ribonucleoprotein complex by acrylamide gel electrophoresis. The results suggest that the polypeptide composition and the products of the cell-associated RNA polymerase are similar to those of the RNA transcriptase associated with influenza virus particles.
TL;DR: A kinetic analysis of the hybridization of a pulse-labeled polysomal mRNA fraction and a 3H-complementary DNA copy of ovalbumin mRNA with an excess of chick DNA suggests that ovalbum in mRNA may be coded by as few as a single gene copy.
TL;DR: The results are consistent with the hypothesis that Ara-C differentially affects the activity of structural and regulatory genes involved in the regulation of GS levels in the retina: Ara- C allows transcription of the enzyme-specific templates, but reversibly inhibits the expression of regulatory genes which limit the accumulation of GS.
Abstract: The analogue of cytidine, cytosine arabinoside (Ara-C), elicited a significant increase in the level of glutamine synthetase (GS) in embryonic chick neural retina in the absence of the steroid inducer of the enzyme. The increase was due to de novo synthesis of GS and was mediated by RNA which accumulated in the presence of the effective concentration of Ara-C. Accumulation of GS did not result from the inhibition of DNA synthesis for which Ara-C is best known. This new effect of Ara-C involves differential suppression of macromolecular synthesis in this system: the concentration of Ara-C which caused maximum GS accumulation suppressed overall protein and RNA syntheses 65–75% without inhibiting the transcription and translation of templates essential for GS synthesis. Withdrawal of Ara-C resulted in restoration of RNA synthesis and cessation of GS accumulation, even though preformed templates for the enzyme were present; however, if all RNA synthesis was arrested with actinomycin D at the time of Ara-C withdrawal, GS continued to accumulate. The results are consistent with the hypothesis that Ara-C differentially affects the activity of structural and regulatory genes involved in the regulation of GS levels in the retina: Ara-C allows transcription of the enzyme-specific templates, but reversibly inhibits the expression of regulatory genes which limit the accumulation of GS.
TL;DR: Kinetics of the appearance of T7-specific RNA polymerase, one of the early gene products, during normal T7 infection show that the capacity of the cells to produce the enzyme decays very rapidly when early mRNA synthesis is terminated by rifampicin or by a natural mechanism programmed by T7.
TL;DR: Results indicate a differential gene expression correlated with rejection or acceptance of the pollentubes by the style, which has a molecular weight different from r-RNA and t- RNA and could be messenger-like of nature.
Abstract: Styles of Petunia hybrida were pollinated (selfed or crossed) and pollentubes were allowed to grow in the presence of 3H-orotic acid. After 24 h RNA was extracted using the cold phenol method and separated on a tandem polyacrylamide gel. Results show that: 1. the synthesis of RNA is more enhanced after cross- than after selfpollination and 2. a restricted number of RNA peaks is responsible for this difference; this RNA has a molecular weight different from r-RNA and t-RNA and could be messenger-like of nature. Results indicate a differential gene expression correlated with rejection or acceptance of the pollentubes by the style.
TL;DR: A complete switch of T7 gene expression from early to late involves at least three different control mechanisms, and it seems that a T7-coded translational repressor may also play a role in the elimination of host functions.
Abstract: BACTERIOPHAGE T7 gene expression is under a unique transcriptional control. The ‘early to late’ switch in T7 development involves a switch from the host Escherichia coli RNA polymerase to the T7-coded enzyme (T7-specific RNA polymerase1), transcribing early mRNA and late mRNA, respectively, from two different regions of the T7 genome2. In addition to the gene 1 coding for T7-specific RNA polymerase, this early–late switch requires the function of another early gene of T7, gene 0.7 or the ‘host shut-off’ gene2,3, to shut off the synthesis of host RNA and T7 early mRNA. This shut-off function is assumed to be an inactivation of the host RNA polymerase, although this has not been proven4,5. Furthermore, we have previously shown that T7 early mRNAs are functionally unstable, although they remain chemically stable late in T7 infection6,7. Thus, it seems that a complete switch of T7 gene expression from early to late involves at least three different control mechanisms. It has been shown that a T7-coded translational repressor may also play a role in the elimination of host functions8.
TL;DR: IIT: Intra-cellular information transfer; mRNA: messenger RNA; pre-mRNA: precursor to mRNA (used synonymously with the formerly used terms), RNP: ribonucleoprotein complex; mRNP: RNP containing mRNA.
Abstract: IIT: Intra-cellular information transfer; mRNA: messenger RNA; pre-mRNA: precursor to mRNA (used synonymously with the formerly used terms, messenger-like RNA (m1RNA), DNA-like RNA (D-RNA) and heterogeneous nuclear RNA (HnRNA); RNP: ribonucleoprotein complex; mRNP: RNP containing mRNA.
TL;DR: Globin mRNA from chick red blood cells at various stages of embryonic development have been isolated and characterized physically and functionally by translation in a cell-free system.
TL;DR: The recent use of a developmental approach to the mechanism of action of estradio1–l7β provides a point of entry which complements and extends the current efforts through the study of receptor proteins and the control of RNA polymerase.
Abstract: Hormonal model systems play an important role in the study of the control of protein synthesis (1) and of development (2). Thus the stimulation of nucleic acid and protein synthesis (3–5) in the immature rat uterus by treatment with estradiol–l7β has received considerable attention. Our recent use of a developmental approach (6–9) to the mechanism of action of estradio1–l7β provides a point of entry which complements and extends the current efforts through the study of receptor proteins and the control of RNA polymerase (10–12).