A 700-bp fragment of the human antithrombin III promoter is sufficient to confer high, tissue-specific expression on human apolipoprotein A-II in transgenic mice

Tremp, Günter; Duchange, Nathalie; Branellec, Didier; Cereghini, Silvia; Tailleux, Anne; Berthou, Laurence; Fiévet, Catherine; Touchet, Nathalie; Schombert, Brigitte; Fruchart, Jean‐Charles; Zakin, Mario M.; Denèfle, Patrice

doi:10.1016/0378-1119(95)00010-4

Cited by 16 publications

(13 citation statements)

References 22 publications

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“…Only differences in protection strength, upstream boundary, or the sequence around the transcription start site in the upper strand could be detected. Our results are somewhat similar to the regulatory features of a transgene AT Ϫ680/ϩ24 nt upstream sequences linked to the apolipoprotein A-II gene (24). In the AT region, four protected areas (I-IV, corresponding to Ϫ138/Ϫ123, Ϫ112/Ϫ104, Ϫ89/Ϫ68, and Ϫ48/Ϫ22 nt, respectively, in the upper strand) were identified by footprint analysis using mouse liver extracts.…”

Section: Discussionsupporting

confidence: 69%

“…We did not observe protection of the putative initiator location (element IV at Ϫ48/Ϫ22 nt in Ref. 24) with extract from any cell line.…”

Section: Discussionmentioning

confidence: 90%

“…HNF4 bound to elements A and C in extracts of HepG2 cells, but not COS1 or HeLa extracts. Interactions of HNF4 with area II of the Ϫ680/ϩ24 nt AT sequences (equivalent to element A) have also been reported in the transgenic (24). However, the absence of HNF4 in CV1-derived cells (COS1 and COS7) has been well documented (25).…”

Section: Discussionmentioning

confidence: 99%

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Regions Flanking Exon 1 Regulate Constitutive Expression of the Human Antithrombin Gene

Fernandez-Rachubinski

Weiner

Blajchman

1996

Journal of Biological Chemistry

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We have identified cis-acting elements and trans-acting factors that regulate constitutive expression of the human antithrombin gene. The activity of the sequences flanking the first exon of the gene was investigated using a luciferase-based reporter assay in transiently transfected HepG2, COS1, BSC40, and HeLa cells. Deletion analysis allowed the mapping of two elements able to promote antithrombin gene transcription in HepG2 and COS1 cells. The first element is located upstream of the first exon (؊150/؉68 nucleotides). The second element is in the first intervening sequence (؉300/؉700 nucleotides) and functions in an orientation opposite to that of the first. Footprint analysis showed three protected areas in the 5 upstream element at ؊92/؊68 (element A), ؊14/؉37 (element B), and ؊126/؊100 nucleotides (element C). These elements acted as enhancers in luciferase reporter assays. Gel retardation analysis demonstrated that two liver-enriched transcription factors, hepatocyte nuclear factor 4 (HNF4) and CCAAT enhancer-binding protein (C/EBPa), bound to the 5 upstream element. HNF4 bound to elements A and C, whereas C/EBPa bound to element B. Element A also interacted with the ubiquitous nuclear hormone receptors chicken ovalbumin upstream promoter transcription factor 1 (COUP-TF1), thyroid hormone receptor ␣ (TR␣), peroxisome proliferator-activated receptor ␣(PPAR␣), and retinoid X receptor ␣ (RXR␣). In HepG2 and BSC40 cells, HNF4, C/EBP␣, and RXR␣ activated luciferase expression from a reporter construct containing the 5-upstream minimal antithrombin gene promoter, while COUP-TF1, TR␣, and HNF3 (␣ or ␤) repressed such expression. Our results show that constitutive expression of the human antithrombin gene depends in part upon the interplay of these transcription factors and suggest that signaling pathways regulated by these factors can modulate antithrombin gene transcription. Human antithrombin (AT)1 is a major inhibitor of a number of serine esterases implicated in blood coagulation (1, 2). The AT gene contains 7 exons and 6 intervening sequences (IVS) in a 14-kilobase pair region of the long arm of chromosome 1. The mechanisms underlying AT gene expression are not well known. The AT gene is expressed primarily in the liver, with lower levels in the kidney and brain (2). AT gene expression is believed to be constitutive, but developmental and hormonal factors are known to influence AT levels in plasma (2). At the transcriptional level, an early report by Prochownik described enhancer activity of a Ϫ340/ϩ1200 nucleotide (nt) fragment of the AT gene (3), while Ochoa et al. (4), investigating the regulation of expression of the human transferrin gene, identified a sequence at Ϫ480/Ϫ458 nt that contained the core motif TCT-TTGACCT. This element, named TF-DRI, was shown to be homologous with an element at Ϫ89/Ϫ75 nt in the human AT gene, and generated shifts in liver nuclear extracts in a tissuespecific manner (4). These observations prompted us to characterize in greater detail elements in this area of the human AT...

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Section: Discussionsupporting

confidence: 69%

“…We did not observe protection of the putative initiator location (element IV at Ϫ48/Ϫ22 nt in Ref. 24) with extract from any cell line.…”

Section: Discussionmentioning

confidence: 90%

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Regions Flanking Exon 1 Regulate Constitutive Expression of the Human Antithrombin Gene

Fernandez-Rachubinski

Weiner

Blajchman

1996

Journal of Biological Chemistry

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“…It has previously been shown that this promoter/enhancer gives transgene expression in liver (Bennoun et al, 1998;Dubois et al, 1991) and kidney (Tremp et al, 1995). Three independent AT3-p53 lines were established (designated lines A, B and C).…”

Section: Generation Of Transgenic Mice Overexpressing P53 In the Livermentioning

confidence: 99%

The consequence of p53 overexpression for liver tumor development and the response of transformed murine hepatocytes to genotoxic agents

et al. 2000

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To analyse the eect of p53 on liver tumor development, we generated transgenic mice overexpressing wild-type p53 in the liver and crossed them with transgenic mice in which the expression of the SV40 large T antigen (TAg) induces hepatic tumors. Remarkably, whereas preneoplastic TAg liver exhibited anisocaryosis and anisocytosis, TAg/p53 liver never presented any dysplastic cells. Moreover, whereas expression of p53 did not aect hepatic development, its constitutive expression in tumorigenic livers resulted in a signi®cantly enhanced apoptosis once nodules had appeared. In contrast, p53 overexpression did not modify the elevated proliferation of TAg-transformed hepatocytes and had no eect on hepatocarcinoma progression. In vitro analysis of primary hepatocytes exposed to various genotoxic agents showed that p53 failed to sensitize normal or TAgtransformed hepatocytes to apoptosis, except when high doses of doxorubicin, UV-B and UV-C radiation were used. Our results con®rmed that the hepatocyte cell type is very resistant to genotoxic agents and showed that constitutive expression of p53 failed to improve their responsiveness. In addition, our results showed that suppression of dysplastic cells, probably by restoring normal cytokinesis and karyokinesis, and enhancement of apoptosis by means of p53 overexpression were insucient to counteract or delay the TAg-induced liver tumoral progression.

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“…Fasting plasma samples from mice were collected into EDTA-containing tubes. Four distinct mouse lines were used in the present study: C57BL/6 control mice, transgenic mice AI-2 expressing human apoA-I (HuAITg mice) (24), transgenic mice AI-2/AIIFu-9 expressing both human apoA-I and human apoA-II (HuAIAIITg mice) (25), and transgenic mice phAT-III/hapoA-II expressing human apoA-II (HuAIITg mice) (34).…”

Section: Methodsmentioning

confidence: 99%

Differential Interaction of the Human Cholesteryl Ester Transfer Protein with Plasma High Density Lipoproteins (HDLs) from Humans, Control Mice, and Transgenic Mice to Human HDL Apolipoproteins

Masson¹,

Duverger²,

Emmanuel³

et al. 1997

Journal of Biological Chemistry

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Plasma high density lipoproteins (HDLs) from humans, from transgenic mice to human apolipoprotein A-I (HuAITg mice), from transgenic mice to human apolipoprotein A-II (HuAIITg mice), from transgenic mice to human apolipoproteins A-I and A-II (HuAIAIITg mice), and from C57BL/6 control mice were isolated, and their ability to interact with the human cholesteryl ester transfer protein (CETP) was studied. Whereas cholesteryl ester transfer rates were gradually enhanced by the addition of moderate amounts of HDL from the different sources, striking differences appeared when HDL levels kept increasing beyond a maximal transfer value. Indeed, while a plateau value corresponding to maximal CETP activity was maintained when raising the concentration of HuAITg HDL and HuAIAIITg HDL, inhibitions could be observed with the highest levels of human, control mouse, and HuAIITg mouse HDL. The concentration-dependent inhibition of CETP activity could be reproduced by the addition of delipidated HDL apolipoproteins from control mice, but it was abolished by a 1-h preheating treatment at 56°C. In contrast, no significant inhibition of CETP activity was observed with the delipidated protein moiety of HuAITg HDL, and cholesteryl ester transfer rates remained unchanged before and after a 1-h, 56°C preheating step. Finally, the CETPmediated transfer of radiolabeled cholesteryl esters from human low density lipoprotein to human HDL was significantly higher in the presence of lipoprotein-deficient plasma from HuAITg mice than in the presence of lipoprotein-deficient plasma from control mice. Interestingly, cholesteryl ester transfer rates measured with both control and HuAITg lipoprotein-deficient plasmas became remarkably similar following a 1-h, 56°C preheating treatment.It is concluded that human, control mouse, and HuAIITg mouse HDL contain a heat-labile lipid transfer inhibitory activity that is absent from HDL of HuAITg and HuAIAIITg mice. Alterations in CETP-lipoprotein binding did not account for differential lipid transfer inhibitory activities.In human plasma, the cholesteryl ester transfer protein (CETP) 1 promotes the exchange of cholesteryl esters and triglycerides between various lipoprotein fractions (1). In vivo, CETP activity results in the net mass transfer of cholesteryl esters from the antiatherogenic high density lipoproteins (HDLs) toward the proatherogenic apoB-containing lipoproteins, i.e. very low density lipoproteins and low density lipoproteins (LDLs) (1). The mechanism of action of human CETP has now been clearly established, and it involves two main steps. In a first step, positively charged amino acids of CETP can interact with negative charges of lipoprotein particles from which, in a second step, it picks up or deposits neutral lipid molecules (for a review, see Ref.2). The CETP-lipoprotein binding has been shown to constitute one major determinant of the CETPmediated neutral lipid transfer reaction (3), and both insufficient and excessive binding of CETP to lipoproteins resulting from alterations in th...

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A 700-bp fragment of the human antithrombin III promoter is sufficient to confer high, tissue-specific expression on human apolipoprotein A-II in transgenic mice

Cited by 16 publications

References 22 publications

Regions Flanking Exon 1 Regulate Constitutive Expression of the Human Antithrombin Gene

Regions Flanking Exon 1 Regulate Constitutive Expression of the Human Antithrombin Gene

The consequence of p53 overexpression for liver tumor development and the response of transformed murine hepatocytes to genotoxic agents

Differential Interaction of the Human Cholesteryl Ester Transfer Protein with Plasma High Density Lipoproteins (HDLs) from Humans, Control Mice, and Transgenic Mice to Human HDL Apolipoproteins

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