Detection of low-level gene induction using in vitro transcription heat shock genes

Love, J.; Vivino, Alfred A.; Minton, Kenneth W.

doi:10.1016/0735-0651(85)90005-6

Cited by 15 publications

(9 citation statements)

References 8 publications

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“…Our understanding of pausing as well as numerous other transcriptioncoupled phenomena has been greatly enhanced through the use of nuclear-run on approaches (Love et al 1985). More recently, Global and Precision Run-On sequencing (GRO/PRO-seq) have provided genome-wide views of the distribution of engaged RNA Polymerases with strand specificity in metazoan organisms (Core et al 2008;Kwak et al 2013).…”

mentioning

confidence: 99%

Divergence of a conserved elongation factor and transcription regulation in budding and fission yeast

Booth¹,

Wang²,

Cheung³

et al. 2016

Genome Res.

146

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Complex regulation of gene expression in mammals has evolved from simpler eukaryotic systems, yet the mechanistic features of this evolution remain elusive. Here, we compared the transcriptional landscapes of the distantly related budding and fission yeast. We adapted the Precision Run-On sequencing (PRO-seq) approach to map the positions of RNA polymerase active sites genome-wide in Schizosaccharomyces pombe and Saccharomyces cerevisiae. Additionally, we mapped preferred sites of transcription initiation in each organism using PRO-cap. Unexpectedly, we identify a pause in early elongation, specific to S. pombe, that requires the conserved elongation factor subunit Spt4 and resembles promoter-proximal pausing in metazoans. PRO-seq profiles in strains lacking Spt4 reveal globally elevated levels of transcribing RNA Polymerase II (Pol II) within genes in both species. Messenger RNA abundance, however, does not reflect the increases in Pol II density, indicating a global reduction in elongation rate. Together, our results provide the first base-pair resolution map of transcription elongation in S. pombe and identify divergent roles for Spt4 in controlling elongation in budding and fission yeast.

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mentioning

confidence: 99%

Divergence of a conserved elongation factor and transcription regulation in budding and fission yeast

Booth¹,

Wang²,

Cheung³

et al. 2016

Genome Res.

146

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“…The expression of hsps induced by a heat shock was also shown to vary strikingly in different media [26]. The effect of conditioned medium could also reside at the transcription level [13]. Its mode of action remains unclear.…”

Section: Activation Of Transcription Of Actin and Hsp Genesmentioning

confidence: 99%

“…A limited number of hsps (5 out of 13) were induced by H 2 0 2 in Salmonella [27] and one only, hsp70, in Chinese hamster ovary cells [28]. On the other hand the whole set of hsp genes was activated by H z 0 2 in Drosophila S2 cells [13]. These discrepancies might be explained by differing sensitivities of the cell lines to H 2 0 2 toxicity, related to differing catalase activities of these cells, resulting in a dose-dependent induction in various cell lines.…”

Section: Activation Of Transcription Of Actin and Hsp Genesmentioning

confidence: 99%

Hydrogen peroxide (H₂O₂) induces actin and some heat‐shock proteins in Drosophila cells

Courgeon

Rollet

Becker

et al. 1988

European Journal of Biochemistry

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Drosophila cells of a clone derived from line Kc were treated with various concentrations of hydrogen peroxide (H202). The concentration of 10 mM was lethal, whereas concentrations of 1 -100 pM did not affect cell viability, rate of multiplication or protein synthesis. The intermediate concentration of 1 mM H z 0 2 was used to study the response of the cells to an oxidative stress.We observed a transitory decrease of the global protein synthesis, which was accompanied by changes in the polypeptide pattern. There was a 2.5-fold increase of the synthesis of the heat-shock proteins 70-68 and 23. The most prominent response was a 6.5-fold increase of actin synthesis 3 h after a 1 mM H 2 0 2 treatment. When aminotriazole (an inhibitor of catalase) was added in association with H202, the increase of actin synthesis became 8.5-fold. Experiments in which catalase was added at various times after H 2 0 2 showed that a 10-min treatment with H 2 0 2 was sufficient to induce actin and heat-shock protein synthesis 3 h later. HzOz was shown to induce the transcriptional activation of an actin gene and of the heat-shock protein genes 70 and 23 within minutes.These results are coherent with the hypothesis that the byproducts of O2 reduction (the superoxide ion and hydrogen peroxide) could be inducers of the heat-shock response. Whether the increase of actin synthesis is a stress-related response, and the mode of action of H 2 0 2 are discussed The heat-shock proteins (hsps), which are synthesised when all living organisms or in vitro cells are exposed to temperatures approximately 5 -20 "C above their optimal growth temperature, can also be induced by a wide range of various agents (for reviews see [l, 21). For example, in Drosophila Kc cells, hsps were shown to be induced without elevation of temperature, by ethanol, arsenite [3], recovery from anoxia [4] and cadmium [5]. The mechanism of induction of hsps is not entirely understood. A heat-shock transcription factor, which binds a consensus sequence in all the heatshock genes has recently been isolated and characterised (for a review see [6]). The heat-shock transcription factor is activated in heat-shocked cells, but what triggers this activation is not known. An interesting question, considering the different agents capable of inducing the synthesis of hsps, is to what extent they might act through a common pathway.We showed previously that in Kc Drosophifa cells the hsps were induced when the cells were reoxygenated after a period of anoxia. At the same time the rate of oxygen consumption was twice the normal rate [4]. This rapid increase of O2 uptake could lead to an overproduction of the products of O2 reduction such as the superoxide ion (0;) and hydrogen peroxide (H20z) [7]. These highly reactive and toxic O2 species were shown to be implicated in many disease states and particularly in tumor promotion [7 -101. We suggested that they might play a role in the chain of events leading to hsp synthesis [4]. In order to test this hypothesis we studied the effect of hydrogen...

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“…Figure 2 shows the accumulation of a transcript complementary to genomic clone pS8 after UV irradiation of 20 J/m2 ( ton, Mass.). Hybridization conditions were as previously described (9,10). Densitometric measurements (GS 300 Scanning Densitometer; Hoefer Scientific Instruments, San Francisco, Calif.) showed enhancement of up to eightfold 5 to 8 h after irradiation and a return to baseline values by 24 h. Calibration with a range of RNA markers from 0.3 to 9 kilobases (kb) indicated that the transcript was 1.0 kb in size.…”

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confidence: 99%

“…The amount of radiolabeled run-on transcript generated by this procedure is proportional to the number of preinitiated transcripts on a given gene. The quantity of labeled transcript is assessed via hybridization against a blot containing the gene(s) of interest (9,10). The in vitro nuclear run-on transcription procedure, hybridization conditions, and subclones of the seven major heat-shock genes have been previously described (10 (4,5) or the hsp23 gene (10).…”

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confidence: 99%

A DNA damage-responsive Drosophila melanogaster gene is also induced by heat shock.

Aa¹,

Smith²,

Kw³

1986

Mol. Cell. Biol.

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A gene isolated by screening Drosophila melanogaster tissue culture cells for DNA damage regulation was also found to be regulated by heat shock. After UV irradiation or heat shock, induction is at the transcriptional level and results in the accumulation of a 1.0-kilobase polyadenylated transcript. The restriction map of the clone bears no resemblance to the known heat shock genes, which are shown to be uninduced by UV irradiation.

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Detection of low-level gene induction using in vitro transcription heat shock genes

Cited by 15 publications

References 8 publications

Divergence of a conserved elongation factor and transcription regulation in budding and fission yeast

Divergence of a conserved elongation factor and transcription regulation in budding and fission yeast

Hydrogen peroxide (H₂O₂) induces actin and some heat‐shock proteins in Drosophila cells

A DNA damage-responsive Drosophila melanogaster gene is also induced by heat shock.

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Detection of low-level gene induction using in vitro transcription heat shock genes

Cited by 15 publications

References 8 publications

Divergence of a conserved elongation factor and transcription regulation in budding and fission yeast

Divergence of a conserved elongation factor and transcription regulation in budding and fission yeast

Hydrogen peroxide (H2O2) induces actin and some heat‐shock proteins in Drosophila cells

A DNA damage-responsive Drosophila melanogaster gene is also induced by heat shock.

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Hydrogen peroxide (H₂O₂) induces actin and some heat‐shock proteins in Drosophila cells