Showing papers on "RNA published in 1975"
TL;DR: Complementarity relationships between this sequence and a purine-rich tract in the ribosome binding site of different bacterial mRNAs suggest that the 3′-end of 16S RNA determines the intrinsic capacity of ribosomes to translate a particular cistron.
Abstract: The sequence of the 3'-terminus of 16S RNA from different bacteria has been determined. Complementarity relationships between this sequence and a purine-rich tract in the ribosome binding site of different bacterial mRNAs suggest that the 3'-end of 16S RNA determines the intrinsic capacity of ribosomes to translate a particular cistron.
1,323 citations
TL;DR: Analysis of the double-stranded, S 1 -resistant fragments indicates that there is a direct relationship between the hairpin loops seen in the electron microscope and secondary structure in aqueous solution.
1,231 citations
TL;DR: The previous finding of similar sequences of methylated nucleotides in mRNA synthesized in vitro by enzymes associated with virus cores indicates that blocked 5' termini may be a characteristic feature of mRNAs that function in eucaryotic cells.
595 citations
TL;DR: Electrophoresis on 9% polyacrylamide gels reveals a dissociable complex containing a pure RNA-RNA hybird that strongly supports the hypothesis that during the initiation step of protein biosynthesis the 3' end of 16S RNA base pairs with the polypurine stretch common to initiator regions in E. coli and bacteriophage mRNAs.
Abstract: Initiation complexes formed by E. coli ribosomes in the presence of 32P-labeled A protein initiator region from R17 bacteriophage Rna have been treated with colicin E3 and disassembled by exposure to 1% sodium dodecyl sulfate. Electrophoresis on 9% polyacrylamide gels reveals a dissociable complex containing the 30-nucleotide-long messenger fragment and the 50-nucleotide-long colicin fragment, which arises from the 3' terminus of the 16S RNA. The complex is a pure RNA-RNA hybird; it is apparently maintained by a seven-base complementarity between the two RNA fragments. Detection of this mRNA-rRNA complex strongly supports the hypothesis that during the initiation step of protein biosynthesis the 3' end of 16S RNA base pairs with the polypurine stretch common to initiator regions in E. coli and bacteriophage mRNAs. The implications of our findings with respect to the molecular mechanism of initiation site selection and mRNA binding to ribosomes, the role of rRNA in ribosome function, and species specificity in translation are explored.
581 citations
TL;DR: Identification of mutant variants of the sequence combined with the in vitro biochemical studies of others has allowed us to tentatively identify the recognition site for each of these proteins, and to suggest how CAP might act at a distance to affect the interaction of RNA polymerase with the promoter.
Abstract: The nucleotide sequence of the lac promoter-operator region has been determined. The 122 base pairs comprising this region include the recognition sites for RNA polymerase, the positive regulatory protein, CAP, and the negative regulatory protein, the repressor. Identification of mutant variants of the sequence combined with the in vitro biochemical studies of others has allowed us to tentatively identify the recognition site for each of these proteins, and to suggest how CAP might act at a distance to affect the interaction of RNA polymerase with the promoter.
480 citations
451 citations
TL;DR: Further studies of mutation in picornaviruses or their RNAs would be fruitful since these viral RNAs are also messenger RNAs and are directly translated, whereas in vivo studies, definitive information is still lacking as to whether alkylation of DNA, RNA, or perhaps protein is the biologically important event.
Abstract: Publisher Summary This chapter discusses the studies relating to the chemical nature of alkylation, from the products of nucleoside alkylation to in vivo effects of alkylating agents that are oncogenic. The reactions of simple methylating agents with nucleosides, nucleotides, and polynucleotides have also been discussed in the chapter. More recent studies of the mechanism of alkylation by nitroso compounds and ethylating agents have shown that both qualitatively and quantitatively the site of alkylation is a function of both the type of reagent used and the conformation and milieu of the nucleic acid. However, translation of this body of knowledge to an understanding of the biological mechanism of alkylation-induced mutagenesis and carcinogenesis is difficult. Further studies of mutation in picornaviruses or their RNAs would be fruitful since these viral RNAs are also messenger RNAs and are directly translated, whereas in vivo studies, definitive information is still lacking as to whether alkylation of DNA, RNA, or perhaps protein, is the biologically important event.
409 citations
TL;DR: Spermatogonia, spermatocytes and Sertoli cells are transcriptionally expressed into heterogeneous nuclear RNA and preribosomal RNA species whereas transcription in sperMatids is predominantly heterogeneousnuclear RNA; and the modification of the chromatin patterns in late spermiogenic steps indicates a stabilized genome that restricts transcriptive functions.
Abstract: A whole-mount electron microscope technique has allowed direct visualization of the transcription process in mouse spermatids. Thes observations have been supported by light and electron microscope autoradiographic techniques that employ [3H]uridine and [3H]arginine in attempts to clarify mechanisms of RNA synthesis and their relationship to nuclear histone changes throughout spermiogenesis. Early spermatid genomes are dispersed almost completely, whereas in later spermiogenic steps the posterior or flagellar nuclear region is readily dispersed and the anterior or subacrosomal nuclear region remains compact. Display of genome segments permits identification of regions where transcription complexes, presumably heterogeneous nuclear RNA species, are seen related to chromatin. These complexes appear as ribonucleoprotein chains, some of them of considerable length, decreasing progressively in number in late spermiogenic steps. This decrease coincides with diminishing rates of [3H]uridine incorporation. Two distinct patterns of chromatin have been identified: a beaded chromatin type associated with transcription complexes encounterd in early spermatids; and a smooth chromatin type not involved in transcriptive activity observed in advanced spermiogenic genomes. Protein particles staining densely with phosphotungstic acid become apparent in nuclei of spermatids after [3H]arginine incorporation becomes significant. There is no structural or autoradiographic evidence for the presence of nucleoli during spermiogenesis. From these data and from previous experimental findings, we conclude that: (a) spermatogonia, spermatocytes and Sertoli cells are transcriptionally expressed into heterogeneous nuclear RNA and preribosomal RNA species whereas transcription in spermatids is predominantly heterogeneous nuclear RNA; and (b) the modification of the chromatin patterns in late spermiogenic steps indicates a stabilized genome that restricts transcriptive functions.
371 citations
TL;DR: Results of the polymerase assays indicate that both L and NS proteins are required to reconstitute transcription activity with a highly purified template composed of only RNA and N protein.
Abstract: Vesicular stomatitis virions, Indiana serotype, were solubilized with high salt solubilizer and separated by ultracentrifugation into a supernatant fraction containing L, G, NS, and M proteins and pellet fraction containing the RNA complexed with N protein NS protein was purified from the supernatnat fluid by sequential chromatography on phosphocellulose and diethylaminoethyl cellulose columns The purified NS protein was assayed in a standard transcription system in combination with purified L protein and purified template (pellet fraction) prepared by renografin or CsCl banding Results of the polymerase assays indicate that both L and NS proteins are required to reconstitute transcription activity with a highly purified template composed of only RNA and N protein The NS protein polymerase activity is destroyed by trypsin but withstands 90 C temperatures for 10 min Cytoplasmic NS protein can substitute for virion NS protein in the in vitro transcription assay
346 citations
TL;DR: An analysis of the methylated constituents of L cell mRNA by a combination of chromatographic methods and enzymatic treatments indicates that they comprise both 2'-O-methyl nucleosides and N6-methyl adenine, and/or 1- methyl adenines, and suggests that the 2'O- methyl nucleotides, Ym, are part of an unusual class of sequences forming the 5' terminus of mRNA.
314 citations
TL;DR: Reovirus mRNA synthesized in vitro by the virus-associated RNA polymerase in the presence of S-adenosylmethionine contains blocked, methylated 5'-termini with the structure, m-7G(5'ppp(5')G-MpCp.
Abstract: Reovirus mRNA synthesized in vitro by the virus-associated RNA polymerase in the presence of S-adenosylmethionine contains blocked, methylated 5'-termini with the structure, m-7G(5')ppp(5')G-MpCp. The functional significance and possible mechanism of formation of this novel 5'-5' terminal nucleotide linkage are discussed.
TL;DR: Most of the poly(adenylic acid)-containing RNA isolated from high-temperature polysomes sediments in sucrose gradients and migrates in gels as a rather narrow band is of sufficient size to code for one particular protein that is found to account for more than half of the total synthesis at high temperature.
Abstract: Heat treatment of D. melanogaster tissue culture cells causes drastic changes in the pattern of protein synthesis and the size distribution of polysomes. Like the heat shock puffs on polytene chromosomes which appear while preexisting puffs regress, heat shock proteins appear on gels while the synthesis of preexisting proteins is sharply reduced, and heat-induced polysomes appear on gradients after preexisting polysomes have disappeared. Most of the poly(adenylic acid)-containing RNA isolated from high-temperature polysomes sediments in sucrose gradients and migrates in gels as a rather narrow band. This RNA is of sufficient size to code for one particular protein that is found to account for more than half of the total synthesis at high temperature. The RNA hybridizes in situ mainly at chromosome sub-division 87B, the site of the major heat shock puff.
TL;DR: Studies on the nature and location of the methylated nucleotides in mRNA synthesized in vitro by vaccinia virus particles revealed an unusual 5'-terminal structure, and evidence that the pyrophosphate group is blocked by 7-methylguanosine and that both 2'-O-methyl-adenosine and 2'- O-methylGuanosine occupy penultimate positions was presented.
Abstract: Studies on the nature and location of the methylated nucleotides in mRNA synthesized in vitro by vaccinia virus particles revealed an unusual 5'-terminal structure. Evidence that the pyrophosphate group is blocked by 7-methylguanosine and that both 2'-O-methyl-adenosine and 2'-O-methylguanosine occupy penultimate positions was presented. According to this model, the 5'-termini of vaccinia virus mRNAs are: 7MeG-5'ppp-5'GMepNp and 7MeG-5'AMepNp.
TL;DR: The poly A-containing mRNA of cultured hamster (BHK-21) cells has been examined with regard to methylation status and the nature of the methylated residues was determined by paper chromatography and electrophoresis of acid and alkaline hydrolysates and of T2 RNase digests.
Abstract: The poly A-containing mRNA of cultured hamster (BHK-21) cells has been examined with regard to methylation status. Steady state-labeled mRNA was obtained by incubating cells for 20-22h in the presence of [methyl-3H]-methionine and 32Pi. The degree of methylation of this RNA was 1.8 methyl groups per 1000 nucleotides, or 4-5 methyl groups on the average per molecule. The nature of the methylated residues was determined by paper chromatography and electrophoresis of acid and alkaline hydrolysates, by DEAE cellulose chromatography of alkaline hydrolysates and of T2 RNase digests, and by examining the effect of subjecting samples to "beta-elimination." Approx. half of the methyl groups occurred in standard ("internal") linkage, 10% as m5Cp and 40% as m6Ap residues. The remainder occurred at least for the most part in "blocked" 5'-termini with the presumptive structure m7G(5')ppp(Nm)p.., where Nm was Gm, m6Am, Um, or Cm.
TL;DR: The structure of the initiation (5′) terminus of mRNA transcribed from CPV by the virus-containing RNA polymerase is of particular interest.
Abstract: METHYLATION of a specific nucleotide occurs at the initial stage of transcription of the double-stranded RNA genome in cytoplasmic polyhedrosis virus (CPV) in the presence of the methyl-group donor, S-adenosyl methionine (SAM)1. Methylation of mRNA has been reported recently not only for other viruses2,3 but also for a mouse L-cell4. Since methylation seems to be coupled with the initiation of mRNA synthesis in CPV, the structure of the initiation (5′) terminus of mRNA transcribed from CPV by the virus-containing RNA polymerase is of particular interest.
TL;DR: The results indicate that methylation of viral mRNA is required for translation in vitro and suggest that mRNA methylation occurs at the initiation step of protein synthesis.
Abstract: Methylated reovirus and vesicular stomatitis virus mRNAs, synthesized in vitro in the presence of S-adenosylmethionine by the virion-associated polymerases (RNA nucleotidyltransferases, EC 2.7.7.6), stimulate protein synthesis by wehat germ extracts to a greater extent than unmethylated mRNAs. Addition of S-adenosylmethionine to a cell-free extract programmed with unmethylated mRNA stimulates protein synthesis and results in methylation of the mRNA. An inhibitor of mRNA methylation. S-adenosylhomocysteine, blocks translation of unmethylated, but not of methylated, mRNAs. Aurintricarboxylic acid, which inhibits polypepetide chain initiation, also prevents mRNA methylation by wheat germ extracts. In contrast, sparsomycin, which inhibits polypeptide chain elongation, does not reduce mRNA methylation. The results indicate that methylation of viral mRNA is required for translation in vitro and suggest that mRNA methylation occurs at the initiation step of protein synthesis.
TL;DR: The steady-state growth rate of Saccharomyces cerevisiae was varied by growing the cells in different media and the total amount of ribonucleic acid per cell was found to decrease as a nonlinear function of decreasing growh rate, while the levels of the different ribosomal RNA species were more closely coordinated.
Abstract: The steady-state growth rate of Saccharomyces cerevisiae was varied by growing the cells in different media. The total amount of ribonucleic acid (RNA) per cell was found to decrease as a nonlinear function of decreasing growh rate. The RNA from cells growing in different media was analyzed by polyacrylamide gel electrophoresis. Although the amounts of both ribosomal RNA and transfer RNA decreased with decreasing growth rate, the ratio of ribosomal to transfer RNA was not constant. As the growth rate was reduced the ribosomal RNA fraction decreased slightly, whereas the transfer RNA fraction increased slightly. Thus the levels of ribosomal and transfer RNA were regulated to similar yet different extents. The levels of the different ribosomal RNA species were more closely coordinated. At all growth rates the ribosomal RNAs (including 5S RNA) were present in equimolar amounts. The rate of protein synthesis in yeast cells also decreased with decreasing growth rate. The low rates of protein synthesis did not appear to be due to limiting numbers of ribosomes or transfer RNA molecules.
TL;DR: The results indicate that low levels of dsRNA promote the formation of an inhibitor which may exist in two forms: one that is reversible by high levels ofdsRNA and one which is irreversible.
TL;DR: In this article, the authors investigated the mechanisms by which dexamethasone (a synthetic glucocorticoid) stimulates the production of mouse mammary tumor virus (MMTV) by cell cultures derived from mammary carcinomas of GR mice.
TL;DR: The 5' terminal structure of the mRNA synthesized in vitro by the virion-associated RNA polymerase of vesicular stomatitis virus in the presence of S-adenosyl-L-methione consists of 7-methyl guanosine linked to 2'-O-methyl adenosine through a 5'-5' pyrophosphate bond as m7G(5'ppp(5')A-m-p.
TL;DR: When the cultured Drosophila cells are subjected to heat shock immediately before labeling with 3H-uridine, there is a drastic alteration in the pattern of gene transcription detected by in situ hybridization.
01 Jan 1975
TL;DR: It is concluded that the accumulation of MMTV RNA is a primary response to dexamethasone and that the rate of synthesis of MM TV RNA is probably accelerated by treatment with dexamETHasone.
Abstract: Gordon M. Ringold, Keith R. Yamamoto, Gordon M. Tomkins, J. Michael Bishop, and Harold E. Varmus Departments of Microbiology and Biochemistry and Biophysics University of California San Francisco, California 94143 Summary We have investigated the mechanisms by which dexamethasone (a synthetic glucocorticoid) stimu- lates the production of mouse mammary tumor virus (MMTV) by cell cultures derived from mam- mary carcinomas of GR mice. Treatment of these cells with dexamethasone stimulates a rapid accu- mulation of intracellular virus-specific RNA which is dependent upon RNA synthesis but not upon DNA or protein synthesis. The effect of dexametha- sone is probably mediated by a specific and satura- ble glucocorticoid receptor. We conclude that the accumulation of MMTV RNA is a primary response to dexamethasone and that the rate of synthesis of MMTV RNA is probably accelerated by treatment with dexamethasone. Introduction Steroid hormones have an important role in regulat- ing specific gene expression in metazoan orga- nisms. It is commonly believed that the various steroids function via a unified mechanism (for re- view, see King and Mainwaring, 1974) in which the hormone diffuses into the target cell and becomes tightly associated with a soluble, steroid-specific re- ceptor protein; the complex then binds to DNA- containing nuclear sites (Harris, 1971; Yamamoto and Alberts, 1972) to initiate the biological response. Thus steroids may act as allosteric effec- tors, altering receptor properties to increase the af- finity of the steroid-receptor complex for the nu- clear binding sites (Jensen et al., 1969; Samuels and Tomkins, 1970). Although evidence consistent with this model has been reported, the initial molecular events subse- quent to interaction of the steroid-receptor complex with the nuclear sites remain obscure. In no case has it been unambiguously shown that the first event detected biochemically actually represents the primary biological effect of the hormone. In fact, in several systems, there is evidence that the pri- mary event is not being directly monitored. For example, the induction of specific proteins by glu-
TL;DR: Mode of action of 5-fluorocytosine (5-FC) and mechanisms of resistance to the drug are discussed on the basis of experiments performed with Candida albicans ATCC 26790 and with 50 selected clinical isolates of C.Albicans belonging to serological type A or B and representing various degrees and models of5- FC resistance (sensitivity).
Abstract: Mode of action of 5-fluorocytosine (5-FC) and mechanisms of resistance to the drug are discussed on the basis of experiments performed with Candida albicans ATCC 26790 and with 50 selected clinical isolates of C. albicans belonging to serological type A or B and representing various degrees and models of 5-FC resistance (sensitivity). Incorporation of 5-fluorouridylic acid into RNA appeared as a prerequisite to antifungal activity, although at a given incorporation rate, growth inhibition varied considerably from one strain to the other. The amino acid pool was unbalanced, and there was evidence for disturbance of protein synthesis. These dysfunctions of RNA probably account for growth inhibition and cell death, whereas up to the present, there was no proof of formation of 5-fluorodeoxyuridylic acid nor of subsequent inhibition of thymidylate synthetase. Incorporation of fluorinated pyrimidine into RNA was lower in normally sensitive type B strains than in normally sensitive ones of type A, whereas the frequency of 5-FC-resistant mutants was the same. The two serological types did not differ in the activity of cytosine permease nor in that of cytosine deaminase. Among 29 clinical isolates with 6-FC resistance (or impaired sensitivity) no instance of cytosine permease deficiency was found. Two isolates (belonging to the serological type A) were deficient in cytosine deaminase, whereas the majority was probably deficient in uridine monophosphate pyrophosphorylase or had a surplus of de novo synthesis of pyrimidines. Relative 5-FC resistance was more common than complete resistance.
TL;DR: The large RNase T1-resistant oligonucleotides of the nondefective Rous sarcoma virus and their transformation-defective mutants were completely or partially mapped on the 30 to 40S viral RNAs.
Abstract: The large RNase T1-resistant oligonucleotides of the nondefective (nd) Rous sarcoma virus (RSV): Prague RSV of subgroup B (PR-B), PR-C and B77 of subgroup C; of their transformation-defective (td0 deletion mutants: td PR-B, td PR-C, and td B77; and of replication-defective (rd) RSV(-) were completely or partially mapped on the 30 to 40S viral RNAs. The location of a given oligonucleotide relative to the poly(A) terminus of the viral RNAs was directly deduced from the smallest size of the poly(A)-tagged RNA fragment from which it could be isolated. Identification of distinct oligonucleotides was based on their location in the electrophoretic/chromatographic fingerprint pattern and on analysis of their RNase A-resistant fragments. The following results were obtained. (i) The number of large oligonucleotides per poly(A)-tagged ffagment increased with increasing size of the fragment. This implies that the genetic map is linear and that a given RNase T1-resistant oligonucleotides has, relative to the poly(A) end, the same location on all 30 to 40S RNA subunits of a given 60 to 70S viral RNA complex, (ii) Three sarcoma-specific oligonucleotides were identified in the RNAs of Pr-B, PR-C and B77 by comparison with the RNAs of the corresponding td viruses...
TL;DR: Lewin et al. as discussed by the authors discussed the evidence that relates hnRNA and mRNA, from kinetic data on mRNA production and analysis of the addition of polyadenylic acid to nu-clear RNA, and consider the limitations that can beplaced on the construction of models for a precur-sor-product relationship between hn RNA andmRNA.
TL;DR: It is now apparent that steroids induce de novo synthesis of both messenger RNA (mRNA) and ribosomal RNA (rRNA); the role of the former in directing the synthesis of specific proteins is reasonably clear but that of the latter remains to be elucidated.
TL;DR: Several lines of evidence suggest that globin messenger ribonucleic acid requires "activation" before it can be utilized and that a messenger activation step of protein synthesis initiation is rate-limiting in this cell-free system.
TL;DR: Recent analyses of the sequence components present in hnRNA and mRNA are discussed and how they may be related to each other and to the organization of the genome.
TL;DR: The nucleotide sequence of an RNA primer molecule for initiation of Rous sarcoma virus DNA synthesis in vitro has been determined and it is confirmed that it is tRNA-Trp, the only species of tRNAs that is dectected in chicked cell tRNA.
TL;DR: Results are presented which suggest that the protein synthesis inhibition is a secondary manifestation of the ability of theDrug to inhibit RNA synthesis, and that of RNA synthesis is most likely the second target site for the action of the drug when bacteria are exposed to it in high concentrations.
Abstract: Nalidixic acid was found to be most bactericidal against various species of gram-negative bacteria at 50 to 200 mug/ml. With all species tested, increases in the concentration of nalidixic acid above this range reduced, rather than increased, its bactericidal effect so that, at levels in the region of 400 mug/ml, the drug was relatively bacteriostatic. Therefore, the mode of action of nalidixic acid at various concentrations was investigated. It was found that at the most bactericidal concentration deoxyribonucleic acid synthesis, but no ribonucleic acid (RNA) or protein synthesis, was inhibited. However at higher concentrations, where the drug is least bactericidal, both RNA and protein synthesis were found to be inhibited. Results are presented which suggest that the protein synthesis inhibition is a secondary manifestation of the ability of the drug to inhibit RNA synthesis, and that of RNA synthesis is most likely the second target site for the action of the drug when bacteria are exposed to it in high concentrations. The clinical implications of these findings are discussed.