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Dynamic Enhancer Methylation - A Previously Unrecognized Switch for Tissue-Type Plasminogen Activator Expression.

Journal article
Authors Mia Magnusson
Emma Xuchun Lu
Pia Larsson
Erik Ulfhammer
Niklas Bergh
Helena Carén
Sverker Jern
Published in PloS one
Volume 10
Issue 10
Pages e0141805
ISSN 1932-6203
Publication year 2015
Published at Wallenberg Laboratory
Sahlgrenska Cancer Center
Institute of Biomedicine, Department of Pathology
Institute of Medicine, Department of Molecular and Clinical Medicine
Pages e0141805
Language en
Subject categories Cardiovascular medicine


Tissue-type plasminogen activator (t-PA), which is synthesized in the endothelial cells lining the blood vessel walls, is a key player in the fibrinolytic system protecting the circulation against occluding thrombus formation. Although classical gene regulation has been quite extensively studied in order to understand the mechanisms behind t-PA regulation, epigenetics, including DNA methylation, still is a largely unexplored field. The aim of this study was to establish the methylation pattern in the t-PA promoter and enhancer in non-cultured compared to cultured human umbilical vein endothelial cells (HUVECs), and to simultaneously examine the level of t-PA gene expression. Bisulphite sequencing was used to evaluate the methylation status, and real-time RT-PCR to determine the gene expression level. While the t-PA promoter was stably unmethylated, we surprisingly observed a rapid reduction in the amount of methylation in the enhancer during cell culturing. This demethylation was in strong negative correlation with a pronounced (by a factor of approximately 25) increase in t-PA gene expression levels. In this study, we show that the methylation level in the t-PA enhancer appears to act as a previously unrecognized switch controlling t-PA expression. Our findings, which suggest that DNA methylation is quite dynamic, have implications also for the interpretation of cell culture experiments in general, as well as in a wider biological context.

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