RNA Polymerase from the Fungus <i>Aspergillus nidulans</i>

Stunnenberg, Henk; Wennekes, L.M.J.; Broek, H.W.J. van den

doi:10.1111/j.1432-1033.1979.tb13167.x

Cited by 13 publications

(3 citation statements)

References 40 publications

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“…crassa are impermeable to these inhibitors (Tisdale & DeBusk, 1972) so the lack of inhibition might be a question of impermeability. However, Stunnenberg et al (1979) have shown that even in vitro three RNA polymerases of A. niduldns are largely or completely insensitive to a-amanitin.…”

Section: Examples Of the Effects Of Various Treatments On Nucleic Acimentioning

confidence: 99%

A Method for Monitoring Macromolecular Synthesis in Filamentous Fungi and its Use to Study Control of Nucleic Acid Synthesis in Aspergillus nidulans

Arst¹,

Scazzocchio²

1980

Microbiology

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A method is described for measuring the syntheses of macromolecules in the filamentous fungus Aspergillus nidulans. It involves growth of mycelium on small filter papers in the presence of radioactively labelled L-leucine to monitor total protein synthesis, and hence growth, and following the incorporation of differently labelled D-uridine (to measure nucleic acid synthesis) or L-leucine (to measure protein synthesis) under various conditions such as starvation for a metabolite or the presence of an inhibitor. Comparison of 3H/14C or 14C/3H labelling ratios (depending on the labelling combination used) shows the effects of the treatments on nucleic acid or protein synthesis. The method allows large numbers of measurements to be made and enables economical use of radioisotopes. Results are presented to show that, under the labelling conditions used, uridine is incorporated mainly into stable RNA. The method is used to demonstrate probable stringent control of stable RNA synthesis in A. niduldns and that inhibitors of protein synthesis such as cycloheximide and anisomycin also inhibit stable RNA synthesis. In contrast, starvation for meso-inositol affects nucleic acid synthesis much more strongly than it affects protein synthesis.

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Section: Examples Of the Effects Of Various Treatments On Nucleic Acimentioning

confidence: 99%

A Method for Monitoring Macromolecular Synthesis in Filamentous Fungi and its Use to Study Control of Nucleic Acid Synthesis in Aspergillus nidulans

Arst¹,

Scazzocchio²

1980

Microbiology

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“…DEAE-Sephacel and heparin-Sepharose CL-6B (Pharmacia) were prewashed and equilibrated as described (15). The density gradient of 5-20% sucrose was made up in 500 mM NaCl/50 mM Tris HCI, pH 7.9/ 5 mM MgCI2/1 mM EDTA/0.5 mM dithiothreitol/0.1 mM PhMeSO2F and run for 40 hr at 36,000 rpm in a Beckman SW 40.1 rotor at 4°C.…”

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confidence: 99%

“…To enrich the active component, we fractionated the protein extract by heparin-Sepharose chromatography (14,15). The column was equilibrated with protein storage buffer (100 mM NaCl, final concentration), and the protein fraction was applied onto the column in the same buffer.…”

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Bioassay for components regulating eukaryotic gene expression: a chromosomal factor involved in the generation of histone mRNA 3' termini.

Stunnenberg¹,

Birnstiel²

1982

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

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We have adapted the oocyte injection procedure for the detection of regulatory components involved in the transcription ofa eukaryotic mRNA gene. Injection of the histone gene repeat h22 DNA ofPsammechinu miliaris into the Xenopwa oocyte nucleus results in correct initiation of the histone mRNAs, but readthrough by RNA polymerase occurs at the 3' end of the H3 histone gene (Hentschel, C. C., Probst, E. & Birnstiel, M. L. (1980) Nature (London) 288, 100-102). Coinjection into the oocyte of a chromosomal salt wash fraction derived from sea urchin embryos results in the generation of authentic 3' termini of the histone H3 mRNA. We have partially purified the protein component by column chromatography and density gradient centrifugation. The regulatory factor binds to heparin columns and, hence, has the properties anticipated ofan RNA-or DNA-binding protein. The sedimentation coefficient of the active component was determined to be about 12 S, suggesting a molecular weight of 200,000-250,000.Two general approaches can be used to study the mechanisms controlling gene expression. The first is to identify the regulatory signals by mutation, followed by an analysis ofthe expression of the mutated genes. Another approach is to characterize regulatory factors (proteins) interacting with these DNA signals.For an understanding of the processes of differentiation, the study of mRNA genes transcribed by polymerase II is of particular importance, because it is the protein-coding genes that primarily determine the cell phenotype.There is now evidence that expression of the different sea urchin histone gene variants is under developmental control (reviewed in ref. 1). This regulation appears to occur both at transcription initiation and termination. Control at transcription initiation is suggested because the regulatory pattern can readily be reproduced by in vitro transcription of nuclei isolated from different embryonic stages ofthe sea urchin (2). That transcription termination also may be controlled developmentally can be inferred from a recent report (3) that transcription in the newt oocyte reads through the 3' termini of histone genes, yielding the large transcripts typical for lampbrush chromosomes of this species.Previous frog oocyte injection experiments have shown that the promoter (4, 5) and terminator signals (6) of all the genes of the sea urchin histone DNA clone h22 appear to be recognized by the frog oocyte transcriptional machinery, although with greatly different efficiencies (7). For the sea urchin H2A histone gene, the faithful generation of correct H2A histone mRNA 3' ends is dependent on the presence of a highly conserved inverted DNA repeat that lies immediately upstream of the 3' mRNA terminus 5' A-C-C-A 3' and on the presence of spacer sequences further downstream (6). The behavior of the H3 histone gene in the oocyte injection experiments is exceptional in that transcription initiation, although varying in different batches of the oocytes, usually occurs at a high rate, but transcription termi...

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An improved method for isolation of Heliothis zea DNA-dependent RNA polymerase: Separation and characterization of a form III RNA polymerase activity

Grula

Weaver

1981

Insect Biochemistry

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RNA Polymerase from the Fungus Aspergillus nidulans

Cited by 13 publications

References 40 publications

A Method for Monitoring Macromolecular Synthesis in Filamentous Fungi and its Use to Study Control of Nucleic Acid Synthesis in Aspergillus nidulans

A Method for Monitoring Macromolecular Synthesis in Filamentous Fungi and its Use to Study Control of Nucleic Acid Synthesis in Aspergillus nidulans

Bioassay for components regulating eukaryotic gene expression: a chromosomal factor involved in the generation of histone mRNA 3' termini.

An improved method for isolation of Heliothis zea DNA-dependent RNA polymerase: Separation and characterization of a form III RNA polymerase activity

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