Transcription modulation in vitro of the fibroin gene exerted by a 200-base-pair region upstream from the "TATA" box.

Tsuda, Masashi; Suzuki, Yuzuru

doi:10.1073/pnas.80.24.7442

Cited by 58 publications

(33 citation statements)

References 24 publications

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“…An approximately 40-bp region from -135 to -175 and also the downstream region from -65 were weakly protected by fraction D, with several hypersensitive sites (C.-C. Hui, unpublished results). The position of these protection sites corresponds generally to the two strongly effective transcription regions determined with the deletion mutants [21].…”

Section: Discussionmentioning

confidence: 99%

“…To investigate which fraction contains stimulatory activities for transcription of the fibroin gene, we examined the functions of individual phosphocellulose fractions by combining them with a small amount of HeLa cell extract, which supplies general transcription factors. In crude posterior-silk-gland extract a stimulation of transcription of the fibroin gene was more marked when circular templates, rather than linear ones, were used [21,221. Therefore, the function of each fraction from the phosphocellulose column chromatography step was examined using circular templates and the modified S1 nuclease assay (Fig.…”

Section: Preferential Stimulation Of Fibr O In Gene Transcript Ion Hymentioning

confidence: 99%

“…The upstream region between -860 and -239 does not contribute to transcription enhancement of the fibroin gene in posterior-silk-gland extract [20,21]. Therefore, we analyzed the stimulatory activity of fraction D by assaying the template activities of three 5'-deletion mutants of the fibroin gene: pFb5'A -238, pFb5'A -72 and pFb5'A -31.…”

Section: Preferential Stimulation Of Fibr O In Gene Transcript Ion Hymentioning

confidence: 99%

“…Single-stranded DNA probe flFb38 carries the coding strand of the fibroin gene (-238 to + 510) on its viral strand [25] and M13Ad4, containing the coding strand of Ad2MLP, was constructed by inserting the smaller EcoRI -SalI fragment of pAd5OO [21] in M13mpll [26].…”

Section: Muter Idsmentioning

confidence: 99%

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Factors involved in preferential transcription of the fibroin gene

Takiya

Suzuki

1989

European Journal of Biochemistry

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Transcription factors for class I1 genes from a tissue source, the posterior silk glands of Bombyx mori, were fractionated by stepwise elution on a phosphocellulose column into five fractions: A (0.04 M KCI flow through), A (0.1 M KCI eluate), B (0.3 M KCI eluate), C (0.5 M KCI eluate) and D (1 M KCI eluate). The minimal requirement for reconstruction of accurate initiation of transcription of the fibroin gene, as well as of the adenovirus 2 major late promoter, was the combination of fractions A, B and D, suggesting that transcription factors from B. mori can recognize general signals of the promoters for class 11 genes and that basic transcription factors are conserved even in distantly diverged species of eukaryotes. To detect activities stimulating the transcription governed by the promoter of fibroin gene, each fraction was tested for its function by supplementing a basal amount of HeLa cell extract. When circular templates were used, stimulatory activities specific for the fibroin gene were detected in fraction D. This preferential transcription is composed of at least three activities; the first and the second dependent on the upstream elements and the third dependent on the sequence downstream from the TATA box. However, when linear templates were used the preference for the fibroin gene was apparently lost and transcription activation by fraction D became general.In order to understand the regulatory mechanisms of gene expression during the development of eukaryotes, the molecular anaylysis of transcription initiation is indispensable. Using various in vivo and in vitro transcription assays, multiple cis-acting elements have been identified in the vicinity of a large number of genes. Specific transcription Factors that bind to some of these elements have been identified [I -101 and, in several cases, their interactions have been demonstrated to give a higher transcriptional activity in vitro [2, 3,[9][10][11][12][13]. Although it is known that the stimulatory factor specific for Ad2MLP binds to its upstream element simultaneously with the transcription factor IID and stabilizes the binding of IID to the TATA box region [3], it is still not clear how these genespecific factors activate or repress transcription. To resolve this problem, it is necessary to analyze the effects caused by the interactions in a transcription system, reconstituted with both purified basal components and regulatory factors.The silk gland of Bombyx mori offers a useful system for analysis of the regulatory mechanism of tissue-specific gene transcription [14]. Transcription of the fibroin gene is restricted to the posterior silk gland and is repeatedly switched on and off during embryonic and larval development, probably under hormonal control have developed an in vitro transcription system composed of extracts from the posterior silk gland. In this system the fibroin gene was actively and preferentially transcribed when compared with Ad2MLP or mouse P-globin gene [20]. This preferential transcription is partly due to upstream el...

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Section: Discussionmentioning

confidence: 99%

Section: Preferential Stimulation Of Fibr O In Gene Transcript Ion Hymentioning

confidence: 99%

Section: Preferential Stimulation Of Fibr O In Gene Transcript Ion Hymentioning

confidence: 99%

Section: Muter Idsmentioning

confidence: 99%

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Factors involved in preferential transcription of the fibroin gene

Takiya

Suzuki

1989

European Journal of Biochemistry

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“…We have prepared a homologous whole-cell extract from the posterior silk glands (20), where the fibroin gene is actively transcribed. In this extract a region covering about 200 base pairs (bp) of DNA sequences upstream from the TATA box of the fibroin gene enhances transcription efficiency (20,21). The upstream sequence can function even in an inverted orientation (21), a feature which is analogous to that of the enhancing elements of other genes characterized in vivo (16,23).…”

mentioning

confidence: 99%

Participation of the upstream region of the fibroin gene in the formation of transcription complex in vitro.

Tsuda¹,

Hirose²,

Suzuki³

1986

Mol. Cell. Biol.

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The addition of exogenous histones has an inhibitory effect on fibroin gene transcription in posterior silk gland extracts. The histones probably disturb a process in complex formation, because when transcription complexes were constructed by preincubation of the templates with the extracts, the inhibitory effect of histones was greatly reduced. Transcription of a fibroin gene construct, pFb5'A-238, having the upstream region beyond the TATA box was relatively less inhibited than that of pFb5'A-44 lacking the upstream region. This tendency toward differential inhibition was observed in the silk gland extracts but not in a HeLa cell extract and persisted even after complex formation in the silk gland extracts, suggesting a specific interaction of the upstream region with some factors in the extracts. The complexes formed on pFb5'A-44 are probably more susceptible to the inhibitory effect of histones. On the basis of these results we propose a participation of the upstream region of the fibroin gene in the formation of stable transcription complexes at the promoter through an interaction with specific factors in the silk gland. Since the transcription-enhancing effect via the upstream region is augmented at a high histone/DNA ratio, it may mimic the in vivo situation in which the fibroin gene can be transcribed in the posterior silk gland even in the presence of excess suppressive materials.

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Specific switching on of silent egg protein genes in vitro by an S-100 fraction in isolated nuclei from male Xenopus.

Tata¹,

Baker²

1985

The EMBO Journal

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A soluble post‐nuclear extract (S‐100) specifically switches on the permanently silent vitellogenin genes in male Xenopus liver nuclei in vitro. Using cloned cDNAs to albumin, globin and vitellogenin mRNAs as hybridization probes, and heparin as an inhibitor of initiation of transcription, run‐on assays with nuclei faithfully reproduced the transcriptional status of these genes in vivo. In the absence of heparin, a liver S‐100 extract from female or male Xenopus treated with estradiol, but not from hormonally untreated males, induced the de novo transcription of vitellogenin genes in nuclei from hormonally naive male hepatocytes. The activation of the dormant genes was dependent on pre‐incubation of nuclei with the competent extract for a minimum of 45 min at 24 degrees C before transcription was initiated. The same extract did not alter significantly the transcription of albumin and globin genes. The S‐100 fraction from hormonally stimulated male hepatocytes exhibited a high degree of tissue specificity; it did not activate the transcription of vitellogenin genes in Xenopus erythrocyte and oviduct nuclei, and only slightly in nuclei from Xenopus XTC‐2 cells, while a soluble extract from oviduct only slightly induced their transcription in male hepatocyte nuclei. This is the first demonstration of a hormone‐ and tissue‐specific de novo activation of dormant genes by a soluble cell extract in isolated nuclei and offers a new approach to analyzing steroid hormone action and tissue‐specific transcription factors in the regulation of gene expression.

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Transcription modulation in vitro of the fibroin gene exerted by a 200-base-pair region upstream from the "TATA" box.

Cited by 58 publications

References 24 publications

Factors involved in preferential transcription of the fibroin gene

Factors involved in preferential transcription of the fibroin gene

Participation of the upstream region of the fibroin gene in the formation of transcription complex in vitro.

Specific switching on of silent egg protein genes in vitro by an S-100 fraction in isolated nuclei from male Xenopus.

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