The
            <i>In Vitro</i>
            Synthesis of Avian Myeloblastosis Viral RNA Sequences

Jacquet, Michel; Groner, Yoram; Monroy, Gladys; Hurwitz, Jerard

doi:10.1073/pnas.71.8.3045

Cited by 62 publications

(28 citation statements)

References 21 publications

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“…The direct engagement of RNA polymerase B in transcribing structural genes has been to date reported only for viral mRNAs (28)(29)(30)(31) and for the fibroin mRNA (32). Our results demonstrate that RNA polymerase B transcribes the ovalbumin gene.…”

Section: Methodssupporting

confidence: 61%

Preferential transcription of the ovalbumin gene in isolated hen oviduct nuclei by RNA polymerase B.

Nguyen-Huu¹,

Sippel²,

Hynes³

et al. 1978

Proc. Natl. Acad. Sci. U.S.A.

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The synthesis of ovalbumin mRNA sequences was studied in isolated nuclei from hen oviduct. Two different methods of analysis were used to distinguish in vitro synthesized from preexisting mRNA sequences: (i) Mercurated ribonucleotides were used for in vitro RNA synthesis, and the newly synthesized RNA was purified by chromatography on sulfhydrylagarose and hybridized to radioactive ovalbumin cDNA. (ii) (3HJUTP was used to label the in vitro synthesized RNA. Hybridization to unlabeled mercurated cDNA, RNase A digestion, and subsequent purification of the hybrids on SH-agarose allowed the quantitation of newly synthesized ovalbumin mRNA sequences. Approximately 0.1% of the newly synthesized RNA was identified as ovalbumin RNA by both methods. The synthesis of ovalbumin RNA progressed during the incubation of nuclei and was sensitive to actinomycin D and low concentrations of a-amanitin. The preferential in vitro transcription of the ovalbumin gene (1000-fold over random transcription of the chicken genome) by RNA polymerase B (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) suggests that the specificity of in vivo RNA synthesis is retained in isolated nuclei.The synthesis of the egg white proteins in the tubular gland cell of the chicken oviduct is primarily related to the cellular accumulation of the corresponding mRNAs (1-7). During stimulation with estradiol the amount of ovalbumin mRNA dramatically increases from a few copies to tens of thousands of copies per cell (2, 4, 5-7). Similar results have been found for the induction of ovomucoid and lysozyme mRNA (7). The steroid-mediated induction of specific mRNA molecules in the chicken oviduct may result from altered rates of synthesis and/or degradation. We therefore studied the synthesis rate of the egg white protein mRNAs. Because it was not possible to measure the rate of specific mRNA synthesis in the animals, we chose isolated oviduct nuclei as the cell-free system that might best reflect the transcriptional activity of the intact cell (8). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.

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Section: Methodssupporting

confidence: 61%

Preferential transcription of the ovalbumin gene in isolated hen oviduct nuclei by RNA polymerase B.

Nguyen-Huu¹,

Sippel²,

Hynes³

et al. 1978

Proc. Natl. Acad. Sci. U.S.A.

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“…The presence of virus-specific DNA stably integrated in the genome of these cells, was shown by nucleic acid hybridization [6,7]. Chromatin from these virus transformed systems has been shown to retain its cell-specific template activity [8][9][10]. Cultured cells derived from mammary carcinomas of mice spontaneously release mouse mammary tumor virus (MMTV), an RNA tumor virus which replicates via a DNA intermediate integrated Present address: Institut Pasteur, Ddpartement de Biologic Mol6culaire, 28, Rue du Dr Roux, 75724 Paris Cgdex 15, France into the DNA of the host (reviews [11,12] ).…”

Section: Introductionmentioning

confidence: 99%

In vitro mouse mammary tumor virus transcription from chromatin A system to study the mechanism of action of glucocorticoid hormones

Crépin¹

1977

FEBS Letters

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“…The retroviruses are a group of single-strand RNA viruses that integrate a double-strand DNA copy of their genome into host cell DNA as an obligatory intermediate oftheir replication cycle (1). Although it has been established that cellular RNA polymerase II (pol II) is responsible for transcription of the integrated viral genome (2,3), it remains uncertain whether information necessary for the initiation of viral transcription is encoded by the viral genome or by host DNA adjacent to the site of integration. The discovery that proviruses contain a directly repeated sequence located at either end of the molecule (4)(5)(6)(7)(8) raised the possibility that proviral DNA contains the control elements necessary to function as an independent transcriptional unit.…”

mentioning

confidence: 99%

Specific transcriptional initiation in vitro on murine type C retrovirus promoters.

Ostrowski¹,

Berard²,

Hager³

1981

Proc. Natl. Acad. Sci. U.S.A.

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We have investigated the ability of molecularly cloned murine type C retroviral DNA to direct accurate initiation of RNA synthesis when added to cell-free extracts. Two different cloned proviruses were used. The first was derived from an integrated molecule of AKR murine leukemia virus and contains adjacent host information. The origin of the second was an unintegrated permuted copy of Harvey murine sarcoma virus. We found that the leukemia virus cloned provirus, as predicted by structural considerations, contained two functional RNA polymerase II promoters located in the U3 region present at either end of the molecule. These promoters initiate transcription at equal rates in vitro. We also found that the permuted sarcoma virus clone contained an RNA polymerase II promoter in the U3 region. Removal of viral sequences 49 bases upstream of the in vitro sarcoma virus initiation site by restriction cleavage results in loss of specific transcription, indicating a role for this information in in vitro promotion. The 5' ends ofin vitro and in vivo viral RNA were compared by nuclease mapping techniques and found to be identical. Based on this evidence, we conclude that murine retroviral genomes contain sufficient information to initiate transcription independent of any host information in vitro and that these viral promoters are probably also active in vivo. In addition to the promoter in U3, Harvey murine sarcoma virus contains a second promoter in vitro that initiates nean the 5' boundary of the transformation-specific (src) region ofthe virus. Initiation by this promoter was insensitive to low levels of a-amanitin, and the RNA transcript could be terminated to yield a 340-nucleotide product. The retroviruses are a group of single-strand RNA viruses that integrate a double-strand DNA copy of their genome into host cell DNA as an obligatory intermediate oftheir replication cycle (1). Although it has been established that cellular RNA polymerase II (pol II) is responsible for transcription of the integrated viral genome (2, 3), it remains uncertain whether information necessary for the initiation of viral transcription is encoded by the viral genome or by host DNA adjacent to the site of integration. The discovery that proviruses contain a directly repeated sequence located at either end of the molecule (4-8) raised the possibility that proviral DNA contains the control elements necessary to function as an independent transcriptional unit. Sequence analysis of these DNA repeats (the long terminal repeat or LTR) for several retroviruses (9-12) confirmed that they contain sequences associated with both initiation and termination of eukaryotic pol II-dependent transcription.Recently, cell-free systems that have the ability to accurately initiate transcription on cloned promoters in vitro have been described (13,14). Subsequently, transcription initiation on various genes (15-17), including a-cloned cDNA fragment of an avian retrovirus (18), have been reported. We have studied the initiation of transcription in vit...

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The In Vitro Synthesis of Avian Myeloblastosis Viral RNA Sequences

Cited by 62 publications

References 21 publications

Preferential transcription of the ovalbumin gene in isolated hen oviduct nuclei by RNA polymerase B.

Preferential transcription of the ovalbumin gene in isolated hen oviduct nuclei by RNA polymerase B.

In vitro mouse mammary tumor virus transcription from chromatin A system to study the mechanism of action of glucocorticoid hormones

Specific transcriptional initiation in vitro on murine type C retrovirus promoters.

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