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DOI: 10.1160/TH10-10-0682
Allelic imbalance of tissue-type plasminogen activator (t-PA) gene expression in human brain tissue
Financial support: The present study was supported by the Swedish Research Council (K2011-65X-14605–09–6), the Swedish Heart-Lung Foundation (20100256), grants from the Swedish State (ALFGBG-148861), 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 Per-Olof Ahl Foundation, and the Göteborg Foundation for Neurological Research. We would also like to thank the SWEGENE Göteborg Genomics Core Facility platform, which was funded by a grant from the Knut and Alice Wallenberg Foundation.
Publication History
Received:
27 October 2010
Accepted after minor revision:
10 February 2011
Publication Date:
28 November 2017 (online)
Summary
We have identified a single-nucleotide polymorphism (SNP) in the t-PA enhancer (-7351C>T), which is associated with endothelial t-PA release in vivo. In vitro studies demonstrated that this SNP is functional at the level of transcription. In the brain, t-PA has been implicated in both physiologic and pathophysiologic processes. The aim of the present study was to examine the effect of the t-PA –7351C>T SNP on t-PA gene expression in human brain tissue. Allelic mRNA expression was measured in heterozygous post-mortem brain tissues using quantitative TaqMan genotyping assay. Protein-DNA interactions were assessed using electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). Significantly higher levels of t-PA mRNA were generated from chromosomes that harboured the wild-type –7351C allele, as compared to those generated from the mutant T allele (for the hippocampus, C to T allelic ratio of ~1.3, p=0.010, n=12; and for the cortex, C to T allelic ratio of ~1.2, p=0.017, n=12). EMSA showed reduced neuronal and astrocytic nuclear protein binding affinity to the T allele, and identified Sp1 and Sp3 as the major transcription factors that bound to the –7351 site. ChIP analyses confirmed that Sp1 recognises this site in intact cells. In conclusion, the t-PA –7351C>T SNP affects t-PA gene expression in human brain tissue. This finding might have clinical implications for neurological conditions associated with enhanced t-PA levels, such as in the acute phase of cerebral ischaemia, and also for stroke recovery.
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