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Thromb Haemost 2010; 104(05): 998-1008
DOI: 10.1160/TH10-04-0243
Blood Coagulation, Fibrinolysis and Cellular Haemostasis
Schattauer GmbH

Allele-specific transcription of the PAI-1 gene in human astrocytes

Karin Hultman
1   Institute of Neuroscience and Physiology, Department of Clinical Neuroscience and Rehabilitation, the Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden
2   Department of Clinical Genetics, Sahlgrenska University Hospital, Gothenburg, Sweden
,
Anna Tjärnlund-Wolf
1   Institute of Neuroscience and Physiology, Department of Clinical Neuroscience and Rehabilitation, the Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden
2   Department of Clinical Genetics, Sahlgrenska University Hospital, Gothenburg, Sweden
,
Jacob Odeberg
3   Atherosclerosis Research Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden
,
Per Eriksson
3   Atherosclerosis Research Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden
,
Christina Jern
1   Institute of Neuroscience and Physiology, Department of Clinical Neuroscience and Rehabilitation, the Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden
2   Department of Clinical Genetics, Sahlgrenska University Hospital, Gothenburg, Sweden
› Institutsangaben Financial support: This study was supported by the Swedish Research Council (K2008-65X-14605–06–3, K2009–64X-12660–12–3), the Swedish Heart-Lung Foundation (20070404, 20090541), grants from the Swedish State (ALFGBG-11206), the Yngve Land Foundation, the Swedish Brain Foundation, the John and Brit Wennerström Foundation, the Rune and Ulla Amlöv Foundation, the Edit Jacobson Foundation, the Göteborg Foundation for Neurological Research, and the Sahlgrenska University Hospital Foundation.
Weitere Informationen

Publikationsverlauf

Received: 21. April 2010

Accepted after major revision: 14. Juli 2010

Publikationsdatum:
24. November 2017 (online)

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Summary

The 4G allele of the PAI-1 –675(4G/5G) insertion/deletion promoter polymorphism has been associated with elevated plasma levels of PAI-1 and an increased risk of myocardial infarction. However, this allele has also been associated with a reduced risk of ischaemic stroke. In the brain, PAI-1 is mainly produced by astrocytes, and can reduce the neurotoxic effects exerted by tissue-type plasminogen activator during pathophysiologic conditions. The aim of the present study was to investigate whether the PAI-1 –675(4G/5G) polymorphism and the linked –844A/G polymorphism affect transcriptional activity of the PAI-1 gene in human astrocytes. Haplotype chromatin immunoprecipitation (haploChIP) was used in order to quantify allele-specific promoter activity in heterozygous cells. Protein-DNA interactions were investigated by electrophoretic mobility shift assay (EMSA). A clear allele-specific difference in PAI-1 gene expression was observed in astrocytes, where the haplotype containing the 4G and the –844A alleles was associated with higher transcriptional activity compared to the 5G and –844G-containing haplotype. EMSA revealed an allele-specific binding of nuclear proteins to the 4G/5G site as well as to the –844A/G site. Supershift experiments identified specific binding of the transcription factors Elf-1 and Elk-1 to the –844G allele. The relative impact of the different sites on allele-specific PAI-1 promoter activity remains to be determined. We demonstrate that common polymorphisms within the PAI-1 promoter affect transcriptional activity of the PAI-1 gene in human astrocytes, thus providing a possible molecular genetic mechanism behind the association between PAI-1 promoter variants and ischaemic stroke.

Keywords

Human astrocytes - haploChIP - plasminogen activator inhibitor type 1 - transcriptional activity
 

  • References

  • 1 Ye Z, Liu EH, Higgins JP. et al. Seven haemostatic gene polymorphisms in coronary disease: meta-analysis of 66,155 cases and 91,307 controls. Lancet 2006; 367: 651-658.
    CrossrefPubMedGoogle Scholar
  • 2 Boekholdt SM, Bijsterveld NR, Moons AH. et al. Genetic variation in coagulation and fibrinolytic proteins and their relation with acute myocardial infarction: a systematic review. Circulation 2001; 104: 3063-3068.
    CrossrefPubMedGoogle Scholar
  • 3 Henry M, Chomiki N, Scarabin PY. et al. Five frequent polymorphisms of the PAI-1 gene: lack of association between genotypes, PAI activity, and triglyceride levels in a healthy population. Arterioscler Thromb Vasc Biol 1997; 17: 851-858.
    CrossrefPubMedGoogle Scholar
  • 4 Anderson JL, Muhlestein JB, Habashi J. et al. Lack of association of a common polymorphism of the plasminogen activator inhibitor-1 gene with coronary artery disease and myocardial infarction. J Am Coll Cardiol 1999; 34: 1778-1783.
    CrossrefPubMedGoogle Scholar
  • 5 Grubic N, Stegnar M, Peternel P. et al. A novel G/A and the 4G/5G polymorphism within the promoter of the plasminogen activator inhibitor-1 gene in patients with deep vein thrombosis. Thromb Res 1996; 84: 431-443.
    CrossrefPubMedGoogle Scholar
  • 6 Abboud N, Ghazouani L, Saidi S. et al. Association of PAI-1 4G/5G and –844G/A gene polymorphisms and changes in PAI-1/tissue plasminogen activator levels in myocardial infarction: a case-control study. Genet Test Mol Biomarkers 2010; 14: 23-27.
    CrossrefPubMedGoogle Scholar
  • 7 Verschuur M, Jellema A, Bladbjerg EM. et al. The plasminogen activator inhibitor-1 (PAI-1) promoter haplotype is related to PAI-1 plasma concentrations in lean individuals. Atherosclerosis 2005; 181: 275-284.
    CrossrefPubMedGoogle Scholar
  • 8 Haselbauer A, Haberbosch W, Tillmanns H. et al. The impact of the PAI-1 A((-844))G promoter polymorphism on the risk and extent of coronary heart disease. Thromb Haemost 2002; 88: 697-698.
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 9 Eriksson P, Kallin B, van ’t Hooft FM. et al. Allele-specific increase in basal transcription of the plasminogen-activator inhibitor 1 gene is associated with myocardial infarction. Proc Natl Acad Sci USA 1995; 92: 1851-1855.
    CrossrefPubMedGoogle Scholar
  • 10 Dawson SJ, Wiman B, Hamsten A. et al. The two allele sequences of a common polymorphism in the promoter of the plasminogen activator inhibitor-1 (PAI-1) gene respond differently to interleukin-1 in HepG2 cells. J Biol Chem 1993; 268: 10739-10745.
    PubMedGoogle Scholar
  • 11 McCormack LJ, Semple JI, Stickland MH. et al. The effect of number of days in culture and plasminogen activator inhibitor-1 (PAI-1) 4G/5G genotype on PAI-1 antigen release by cultured human umbilical vein endothelial cells. Thromb Res 1998; 92: 199-206.
    CrossrefPubMedGoogle Scholar
  • 12 Bentley P, Peck G, Smeeth L. et al. Causal relationship of susceptibility genes to ischemic stroke: comparison to ischemic heart disease and biochemical determinants. PLoS One 2010; 05: e9136.
    CrossrefPubMedGoogle Scholar
  • 13 Roest M, van der Schouw YT, Banga JD. et al. Plasminogen activator inhibitor 4G polymorphism is associated with decreased risk of cerebrovascular mortality in older women. Circulation 2000; 101: 67-70.
    CrossrefPubMedGoogle Scholar
  • 14 Gravanis I, Tsirka SE. Tissue plasminogen activator and glial function. Glia 2005; 49: 177-183.
    CrossrefPubMedGoogle Scholar
  • 15 Chmielewska J, Ranby M, Wiman B. Kinetics of the inhibition of plasminogen activators by the plasminogen-activator inhibitor. Evidence for ‘second-site’ interactions. Biochem J 1988; 251: 327-332.
    CrossrefPubMedGoogle Scholar
  • 16 Calabresi P, Napolitano M, Centonze D. et al. Tissue plasminogen activator controls multiple forms of synaptic plasticity and memory. Eur J Neurosci 2000; 12: 1002-1012.
    CrossrefPubMedGoogle Scholar
  • 17 Tsirka SE, Rogove AD, Bugge TH. et al. An extracellular proteolytic cascade promotes neuronal degeneration in the mouse hippocampus. J Neurosci 1997; 17: 543-552.
    CrossrefPubMedGoogle Scholar
  • 18 Yepes M, Sandkvist M, Moore EG. et al. Tissue-type plasminogen activator induces opening of the blood-brain barrier via the LDL receptor-related protein. J Clin Invest 2003; 112: 1533-1540.
    CrossrefPubMedGoogle Scholar
  • 19 Yang D, Nemkul N, Shereen A. et al. Therapeutic administration of plasminogen activator inhibitor-1 prevents hypoxic-ischemic brain injury in newborns. J Neurosci 2009; 29: 8669-8674.
    CrossrefPubMedGoogle Scholar
  • 20 Nagai N, De Mol M, Lijnen HR. et al. Role of plasminogen system components in focal cerebral ischemic infarction: a gene targeting and gene transfer study in mice. Circulation 1999; 99: 2440-2444.
    CrossrefPubMedGoogle Scholar
  • 21 Docagne F, Nicole O, Gabriel C. et al. Smad3-dependent induction of plasminogen activator inhibitor-1 in astrocytes mediates neuroprotective activity of transforming growth factor-beta 1 against NMDA-induced necrosis. Mol Cell Neurosci 2002; 21: 634-644.
    CrossrefPubMedGoogle Scholar
  • 22 Knight JC, Keating BJ, Rockett KA. et al. In vivo characterization of regulatory polymorphisms by allele-specific quantification of RNA polymerase loading. Nat Genet 2003; 33: 469-475.
    CrossrefPubMedGoogle Scholar
  • 23 Weeks JR, Hardin SE, Shen J. et al. Locus-specific variation in phosphorylation state of RNA polymerase II in vivo: correlations with gene activity and transcript processing. Genes Dev 1993; 07: 2329-2344.
    CrossrefPubMedGoogle Scholar
  • 24 Jaffe EA, Nachman RL, Becker CG. et al. Culture of human endothelial cells derived from umbilical veins. Identification by morphologic and immunologic criteria. J Clin Invest 1973; 52: 2745-2756.
    CrossrefPubMedGoogle Scholar
  • 25 Wolf AT, Medcalf RL, Jern C. The t-PA –7351C>T enhancer polymorphism decreases Sp1 and Sp3 protein binding affinity and transcriptional responsiveness to retinoic acid. Blood 2005; 105: 1060-1067.
    PubMedGoogle Scholar
  • 26 Hultman KBF, Nilsson M, Wilhelmsson U. et al. Expression of PAI-1 and PN-1 in human astrocytes; response to injury-related factors. J Neurosci Res 2010; 88: 2441-2449.
    PubMedGoogle Scholar
  • 27 Morange PE, Saut N, Alessi MC. et al. Association of plasminogen activator inhibitor (PAI)-1 (SERPINE1) SNPs with myocardial infarction, plasma PAI-1, and metabolic parameters: the HIFMECH study. Arterioscler Thromb Vasc Biol 2007; 27: 2250-2257.
    CrossrefPubMedGoogle Scholar
  • 28 Buchwalter G, Gross C, Wasylyk B. Ets ternary complex transcription factors. Gene 2004; 324: 1-14.
    CrossrefPubMedGoogle Scholar