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

Karin Hultman; Anna Tjärnlund-Wolf; Jacob Odeberg; Per Eriksson; Christina Jern

doi:10.1160/TH10-04-0243

Thrombosis and Haemostasis, Table of Contents

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

¹Institute of Neuroscience and Physiology, Department of Clinical Neuroscience and Rehabilitation, the Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden

²Department of Clinical Genetics, Sahlgrenska University Hospital, Gothenburg, Sweden

,

Anna Tjärnlund-Wolf

¹Institute of Neuroscience and Physiology, Department of Clinical Neuroscience and Rehabilitation, the Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden

²Department of Clinical Genetics, Sahlgrenska University Hospital, Gothenburg, Sweden

,

Jacob Odeberg

³Atherosclerosis Research Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden

,

Per Eriksson

³Atherosclerosis Research Unit, Department of Medicine, Karolinska Institute, Stockholm, Sweden

,

Christina Jern

¹Institute of Neuroscience and Physiology, Department of Clinical Neuroscience and Rehabilitation, the Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden

²Department of Clinical Genetics, Sahlgrenska University Hospital, Gothenburg, Sweden

› Author Affiliations

Abstract

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

Full Text

References

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