Initiation of Protein S mRNA synthesis in human liver and various cell lines

Wolf, Cornelia; Cupers, R; Bertina, Rogier M.; Vos, Hans L.

doi:10.1111/j.1538-7836.2005.01138.x

Cited by 6 publications

(13 citation statements)

References 17 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The different construct sizes in both studies could cause the different effects of the mutations. As described above, Tatewaki et al (33) used a construct that ranged from Ϫ282 to Ϫ161, thereby lacking several of the previously identified transcription start sites (32), whereas the construct used in this report is larger, spanning nucleotides Ϫ370 to Ϫ1 in relation to the PROS1 translational start codon. Co-transfection of HepG2 cells with FOXA2 and PS370-luc or the PS370-luc FOXA2 mutant resulted in a slight induction of the wild type PS370-luc but not of the FOXA2 mutant construct.…”

Section: Discussionmentioning

confidence: 99%

“…EMSA buffers were purchased from Active Motif (Carlsbad, CA) and used according to the manufacturer's recommendations. Double-stranded oligonucleotides were end-labeled using [␥- 32 P]ATP and T4 polynucleotide kinase. The position numbers for the oligonucleotides in Fig.…”

Section: Methodsmentioning

confidence: 99%

“…The promoter and the first exon are absent from the PROS2 gene, however (29 -31). Transcription from the PROS1 promoter is directed from multiple start sites (32), and recently the PROS1 promoter was shown to contain a forkhead box A2 (FOXA2)-binding site and an Sp1-binding site (33).…”

Section: ) Protein S (Ps)mentioning

confidence: 99%

See 2 more Smart Citations

The Constitutive Expression of Anticoagulant Protein S Is Regulated through Multiple Binding Sites for Sp1 and Sp3 Transcription Factors in the Protein S Gene Promoter

Wolf

Cupers

Bertina

et al. 2006

Journal of Biological Chemistry

View full text Add to dashboard Cite

Protein S (PS) is a vitamin K-dependent plasma protein that inhibits blood coagulation by serving as a nonenzymatic cofactor for activated protein C in the protein C anticoagulant pathway. Low PS levels are a risk factor for the development of deep venous thrombosis. The regulation of PS levels through transcriptional regulation of the PS gene was investigated in this report. A minimal PS gene promoter 370 bp upstream from the translational initiation codon was sufficient for maximal promoter activity in transient transfections regardless of the cell type. A pivotal role for Sp1 in the constitutive expression of the PS gene was demonstrated through electrophoretic mobility shift assay experiments, transient expression of mutant PS promoter-reporter gene constructs, and chromatin immunoprecipitations in HepG2 cells. At least four Sp-binding sites were identified. The two sites most proximal to the translational start codon were found to be indispensable for PS promoter activity, whereas mutation of the two most distal Sp-binding sites had a negligible influence on basal promoter activity. In addition, all other major promoter-binding proteins that were found by electrophoretic mobility shift assay could be positively identified in supershift assays. We identified binding sites for the hepatocyte-specific forkhead transcription factor FOXA2, nuclear factor Y, and the cAMP-response element-binding protein/activating transcription factor family of transcription factors. Their relevance was investigated using site-directed mutagenesis.The coagulation cascade is a complex system in which the consecutive activation of multiple coagulation factors leads to the production of thrombin and ultimately to the formation of fibrin polymers, the primary component of blood clots (for a recent review see Ref. 1). Protein S (PS)2 is a vitamin K-dependent plasma protein that functions as a nonenzymatic cofactor for activated protein C in the down-regulation of the coagulation cascade via proteolytic inactivation of coagulant factors Va and VIIIa (2-5). PS has also been shown to display activated protein C-independent anticoagulant activity in purified systems as well as in plasma (6 -8). Recent studies indicate that PS may have a second function unrelated to coagulation in the clearance of apoptotic cells (9, 10).Over the past two decades low PS plasma levels have become a well established risk factor for the development of deep venous thrombosis (11-13). However, not all mechanisms underlying low plasma PS levels have been fully characterized. Hereditary PS deficiency has been shown to be an autosomal dominant trait, and many causative genetic mutations have been described in the PS gene (14,15). On the other hand, PS deficiency can also be acquired throughout life by conditions such as oral contraceptive use and liver disease (16). To better understand the different functions of PS and the possible causes of PS deficiency, more information is needed on the regulation of the PS gene, mRNA, and protein.The major source of circulati...

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

The Constitutive Expression of Anticoagulant Protein S Is Regulated through Multiple Binding Sites for Sp1 and Sp3 Transcription Factors in the Protein S Gene Promoter

Wolf

Cupers

Bertina

et al. 2006

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…The promoter and first exon are absent from the PROS2 gene, and the promoter region of PROS1 has been poorly investigated in contrast to the coding regions. It has been reported that transcription from the PROS1 promoter is directed from multiple start sites and that the PROS1 5Ј-flanking region lacks the characteristic "CAAT" and "TATA" boxes (23).…”

Section: Protein S (Ps)mentioning

confidence: 99%

Down-regulation of PROS1 Gene Expression by 17β-Estradiol via Estrogen Receptor α (ERα)-Sp1 Interaction Recruiting Receptor-interacting Protein 140 and the Corepressor-HDAC3 Complex

Suzuki

Sanda

Miyawaki

et al. 2010

Journal of Biological Chemistry

View full text Add to dashboard Cite

Pregnant women show a low level of protein S (PS) in plasma,which is known to be a risk for deep venous thrombosis. 17␤-Estradiol (E 2 ), an estrogen that increases in concentration in the late stages of pregnancy, regulates the expression of various genes via the estrogen receptor (ER). Here, we investigated the molecular mechanisms behind the reduction in PS levels caused by E 2 in HepG2-ER␣ cells, which stably express ER␣, and also the genomic ER signaling pathway, which modulates the liganddependent repression of the PS␣ gene (PROS1). We observed that E 2 repressed the production of mRNA and antigen of PS. A luciferase reporter assay revealed that E 2 down-regulated PROS1 promoter activity and that this E 2 -dependent repression disappeared upon the deletion or mutation of two adjacent GCrich motifs in the promoter. An electrophoretic mobility shift assay and DNA pulldown assay revealed that the GC-rich motifs were associated with Sp1, Sp3, and ER␣. In a chromatin immunoprecipitation assay, we found ER␣-Sp protein-promoter interaction involved in the E 2 -dependent repression of PROS1 transcription. Furthermore, we demonstrated that E 2 treatment recruited RIP140 and the NCoR-SMRT-HDAC3 complex to the PROS1 promoter, which hypoacetylated chromatin. Taken together, this suggested that E 2 might repress PROS1 transcription depending upon ER␣-Sp1 recruiting transcriptional repressors in HepG2-ER␣ cells and, consequently, that high levels of E 2 leading to reduced levels of plasma PS would be a risk for deep venous thrombosis in pregnant women.

show abstract

“…Transcription from the PROS1 promoter is directed from multiple start sites (de Wolf et al , 2005), and recently two groups have characterised essential regulatory elements in the 5′‐flanking region of the PROS1 (Tatewaki et al , 2003; de Wolf et al , 2006). Various transcription factor binding sites were identified within the first 400‐bp proximal to the PROS1 translational start site, including multiple binding sites for ubiquitous transcription factors Sp1 and Sp3.…”

mentioning

confidence: 99%

An Sp1 binding site mutation of the PROS1 promoter in a patient with protein S deficiency

et al. 2007

View full text Add to dashboard Cite

Initiation of Protein S mRNA synthesis in human liver and various cell lines

Cited by 6 publications

References 17 publications

The Constitutive Expression of Anticoagulant Protein S Is Regulated through Multiple Binding Sites for Sp1 and Sp3 Transcription Factors in the Protein S Gene Promoter

The Constitutive Expression of Anticoagulant Protein S Is Regulated through Multiple Binding Sites for Sp1 and Sp3 Transcription Factors in the Protein S Gene Promoter

Down-regulation of PROS1 Gene Expression by 17β-Estradiol via Estrogen Receptor α (ERα)-Sp1 Interaction Recruiting Receptor-interacting Protein 140 and the Corepressor-HDAC3 Complex

An Sp1 binding site mutation of the PROS1 promoter in a patient with protein S deficiency

Contact Info

Product

Resources

About