Specific in vitro transcription of conalbumin gene is drastically decreased by single-point mutation in T-A-T-A box homology sequence.

Wasylyk, Bohdan; Derbyshire, R.; Guy, A.; Molko, D.; Roget, André; Téoule, R.; Chambon, Pierre

doi:10.1073/pnas.77.12.7024

Cited by 180 publications

(59 citation statements)

References 23 publications

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“…Similar observations were recently reported by Corden et al (17) for the upstream border of the major late promoter of Ad2. In addition, they demonstrated that point mutations and small deletions in the T-A-T-A sequence drastically reduced transcription (18,19). A requirement for the T-A-T-A sequence in the initiation of specific transcription in vitro has been implicated with varying precision for the chicken conalbumin (17) and ovalbumin (20) genes, and possibly for the simian virus 40 early region (21).…”

Section: Discussionmentioning

confidence: 99%

DNA sequences affecting specific initiation of transcription in vitro from the EIII promoter of adenovirus 2.

Lee¹,

Roeder²,

Wold³

1982

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

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We have identified those sequences affecting the level of specific initiation of transcription in vitro from the EMI promoter of adenovirus 2. Mutants containing deletions in and around the initiation sites were constructed in cloned viral DNA fragments and assayed for their ability to initiate transcription in vitra Three classes of mutants were studied with deletions in the following regions: -38 to -268, -21 to -71 (which includes the T-A-T-A-A box), and -29 through the cap sites (+ l and +3). Deletions that remove some or all of the area from -28 to several nucleotides downstream from the cap sites essentially abolished specific transcription. Small deletions in the region -30 to -41 reduced transcription to -60% ofwild type; larger deletions in the region -35 to -268 reduced transcription to 30-40% ofwild type. Deletions beginning from approximately + 10 to +25 and extending further downstream reduced transcription to 20-40% of wild type, whereas a deletion beginning at +31 had little or no effect. Our results suggest that the region including the

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Section: Discussionmentioning

confidence: 99%

DNA sequences affecting specific initiation of transcription in vitro from the EIII promoter of adenovirus 2.

Lee¹,

Roeder²,

Wold³

1982

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

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“…Therefore, this box appears to be the main promoter element for transcription of the human corticotropinlp-lipotropin precursor gene in aitro. Using a singlebase-pair transversion mutant, Wasylyk et al [37] have shown that the 'TATA box' is essential for transcription of the chicken conalbumin gene in aitro. The requirement for this box in cell-free transcription has also been reported for several other eukaryotic genes [25,38,39].…”

Section: Discussionmentioning

confidence: 99%

Sequence Requirement for Transcription in vitro of the Human Corticotropin/β‐Lipotropin Precursor Gene

Notake

Kurosaki

Yamamoto

et al. 1983

European Journal of Biochemistry

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Using HeLa whole cell extracts, we have demonstrated that transciption in ziitro of the cloned human and bovine corticotropinlj-lipotropin precursor genes is initiated accurately and efficiently. D N A sequences required for promoter function have been assessed by using a series of 5'-deletion mutants of a fusion gene that contains the 5'-flanking sequence and capping site of the human corticotropinlj-lipotropin precursor gene and the structural sequence of the herpes simplex virus thymidine kinase gene. The results obtained have shown that the region between 22 base pairs and 35 base pairs upstream from the capping site is essential for the correct and efficient transcriptional initiation in aitro. Thus, the 'TATA box' present in this region seems to be the main promoter element for transcription of the human corticotropin//?-lipotropin precursor gene in the HeLa cell-free system.We have also developed a transcription system in aitro from the corticotropin-producing mouse pituitary tumor cell line AtT-20 in culture. Deletion mapping of the fusion gene promoter has indicated that the 'TATA box' region is required for the accurate and efficient transcriptional initiation in this system as well. Characteristic ofthis system is that the deletion of the sequence lying between 53 base pairs and 59 base pairs upstream from the capping site increases the transcriptional efficiency. Because this effect is observed in the AtT-20 cell-free system, but hardly in the HeLa cell-free system, it seems reasonable to assume that the interaction of this upstream sequence with some factor(s) in the AtT-20 cell extract is responsible for the modulation of transcription of the human corticotropin/ 0-lipotropin precursor gene.

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“…The pellets were re-suspended in 40ml X-buffer (lmM MgSO4, 0.002% gelatine, 2AsM sodium phosphate buffer pH 7.2) and the phage suspension centrifuged for 16h at 80,000 x g. The resulting pellets were re-suspended in 1-2ml X-buffer. DNA was then isolated by phenol extraction and extensive dialysis against TES-buffer (5C1M NaCl, with filamentous phage (typically less than 10%) 13,17). It seemed to us that a possible explanation for this behaviour could be offered by the study of Radman et al ) on mismatch repair acting on duplex DNA molecules of the X genome containing one or two mismatch positions.…”

Section: Methodsmentioning

confidence: 99%

Directed mutagenesis of DNA cloned in filamentous phage: influence of hemimethylated GATC sites on marker recovery from restriction fragments

1982

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Gapped duplex DNA molecules of recombinant genomes of filamentous phage are constructed in vitro. Denatured restriction fragments covering (part of) the precisely constructed gap are hybridized to the gapped duplex DNA molecules to form ternary duplices. The two strands of the ternary duplex molecules carry different genetic markers within the region spanned by the restriction fragment leading to a one base pair mismatch or to an insertion loop of 93 nucleotides, respectively. The two strands also vary with respect to A-methylation in GATC sites. In cases of asymmetrical methylation, transfection of E.coli with these heteroduplex molecules leads to marker recoveries with a pronounced bias in favour of the marker encoded by the methylated strand. This effect at least partly explains the comparably low marker yields achieved in previous directed mutagenesis experiments using filamentous phage as the vector. The results suggest how these procedures can be optimized. Precise construction of a 93 bp insertion in 9.5% marker yield is described.

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Specific in vitro transcription of conalbumin gene is drastically decreased by single-point mutation in T-A-T-A box homology sequence.

Cited by 180 publications

References 23 publications

DNA sequences affecting specific initiation of transcription in vitro from the EIII promoter of adenovirus 2.

DNA sequences affecting specific initiation of transcription in vitro from the EIII promoter of adenovirus 2.

Sequence Requirement for Transcription in vitro of the Human Corticotropin/β‐Lipotropin Precursor Gene

Directed mutagenesis of DNA cloned in filamentous phage: influence of hemimethylated GATC sites on marker recovery from restriction fragments

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