A Globin Enhancer Acts by Increasing the Proportion of Erythrocytes Expressing a Linked Transgene

Sutherland, Heidi G.; Martin, David I. K.; Whitelaw, Emma

doi:10.1128/mcb.17.3.1607

Cited by 74 publications

(51 citation statements)

References 50 publications

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“…Similar conclusions were drawn from the analysis of cells transfected with reporter constructs with or without the Simian virus (SV)40 enhancer. The presence of the enhancer resulted in a larger proportion of cells expressing the reporter, but not in an increase in expression levels within individual cells (12)(13)(14)(15)(16). These data all provide support for an all-or-none transcriptional activation mechanism.…”

supporting

confidence: 59%

Graded transcriptional response to different concentrations of a single transactivator

Kringstein

Rossi

Hofmann

et al. 1998

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

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Threshold mechanisms of transcriptional activation are thought to be critical for translating continuous gradients of extracellular signals into discrete all-or-none cellular responses, such as mitogenesis and differentiation. Indeed, unequivocal evidence for a graded transcriptional response in which the concentration of inducer directly correlates with the level of gene expression in individual eukaryotic cells is lacking. By using a novel binary tetracycline regulatable retroviral vector system, we observed a graded rather than a threshold mechanism of transcriptional activation in two different model systems. When polyclonal populations of cells were analyzed at the single cell level, a dose-dependent, stepwise increase in expression of the reporter gene, green f luorescent protein (GFP), was observed by f luorescence-activated cell sorting. These data provide evidence that, in addition to the generally observed all-or-none switch, the basal transcription machinery also can respond proportionally to changes in concentration of extracellular inducers and trancriptional activators.

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supporting

confidence: 59%

Graded transcriptional response to different concentrations of a single transactivator

Kringstein

Rossi

Hofmann

et al. 1998

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

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“…We have to date only examined transgene expression at the population level, and it is possible that there is variation in the number of cells in the spleen cell population that express the wt or mutated transgenes as opposed to the absolute levels produced per cell. Such effects have been observed in the ␤-globin gene system as well as for the CD2 gene (47)(48)(49).…”

Section: Discussionmentioning

confidence: 90%

A NF-κB/Sp1 Region Is Essential for Chromatin Remodeling and Correct Transcription of a Human Granulocyte- Macrophage Colony-Stimulating Factor Transgene

Cakouros

Cockerill

Bert

et al. 2001

The Journal of Immunology

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The GM-CSF gene is expressed following activation of T cells. The proximal promoter and an upstream enhancer have previously been characterized using transfection and reporter assays in T cell lines in culture. A 10.5-kb transgene containing the entire human GM-CSF gene has also been shown to display inducible, position-independent, copy number-dependent transcription in mouse splenocytes. To determine the role of individual promoter elements in transgene function, mutations were introduced into the proximal promoter and activity assessed following the generation of transgenic mice. Of four mutations introduced into the transgene promoter, only one, in an NF-κB/Sp1 region, led to decreased induction of the transgene in splenocytes or bone marrow-derived macrophages. This mutation also affected the activity of reporter gene constructs stably transfected into T cell lines in culture, but not when transiently transfected into the same cell lines. The mutation alters the NF-κB family members that bind to the NF-κB site as well as reducing the binding of Sp1 to an adjacent element. A DNase I hypersensitive site that is normally generated at the promoter following T cell activation on the wild-type transgene does not appear in the mutant transgene. These results suggest that the NF-κB/Sp1 region plays a critical role in chromatin remodeling and transcription on the GM-CSF promoter in primary T cells.

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“…However, additional sequences present mainly in the 0.7-kb element C seem to be necessary to maintain a high stable level of transcriptional activity. cisActing positive elements can be seen as sequences that increase the probability of formation of an active chromatin domain rather than as sequences that directly increase the rate of transcription (40,41). In this model, sequences present in element C could act by increasing the number of cells expressing the transgenes in the brain.…”

Section: High Level Expression Of the Aldolase C Gene Requires The Prmentioning

confidence: 99%

Upstream Elements Involved in Vivo in Activation of the Brain-specific Rat Aldolase C Gene

Skala

Porteu

Thomas

et al. 1998

Journal of Biological Chemistry

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The rat aldolase C gene encodes a glycolytic enzyme strongly expressed in adult brain. We previously reported that a 115-base pair (bp) promoter fragment was able to ensure the brain-specific expression of the chloramphenicol acetyltransferase (CAT) reporter gene in transgenic mice, but only at a low level (Thomas, M., Makeh, I., Briand, P., Kahn, A., and Skala, H. (1993) Eur. J. Biochem. 218, 143-151). Here we show that in vivo activation of this promoter at a high level requires cooperation between an upstream 0.6-kilobase pair (kb) fragment and far upstream sequences. In the 0.6-kb region, a 28-bp DNA element is shown to include overlapping in vitro binding sites for POU domain regulatory proteins and for the Winged Helix hepatocyte nuclear factor-3␤ factor. An hepatocyte nuclear factor-3␤-binding site previously described in the short proximal promoter fragment is also shown to interact in vitro with POU proteins, although with a lower affinity than the 28-bp motif. Additional binding sites for POU factors were detected in the upstream 0.6-kb sequences. Progressive deletion in this region resulted in decreased expression levels of the transgenes in mice, suggesting synergistic interactions between these multiple POUbinding sites. We propose that DNA elements characterized by a dual binding specificity for both POU domain and Winged Helix transcription factors could play an essential role in the brain-specific expression of the aldolase C gene and other neuronal genes.

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A Globin Enhancer Acts by Increasing the Proportion of Erythrocytes Expressing a Linked Transgene

Cited by 74 publications

References 50 publications

Graded transcriptional response to different concentrations of a single transactivator

Graded transcriptional response to different concentrations of a single transactivator

A NF-κB/Sp1 Region Is Essential for Chromatin Remodeling and Correct Transcription of a Human Granulocyte- Macrophage Colony-Stimulating Factor Transgene

Upstream Elements Involved in Vivo in Activation of the Brain-specific Rat Aldolase C Gene

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