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Published on: April 9, 2018
Analysis of CD39/ATP diphosphohydrolase (ATPDase) expression in endothelial cells, platelets and leukocytes
K Koziak1, J Sévigny, S C Robson
1Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA.
Insights
Extracellular ATP hydrolysis by ATP diphosphohydrolase (ATPDase), identified as CD39, regulates key cellular processes. CD39 expression correlates with ATPDase activity, influencing thrombotic and inflammatory responses.
Area of Science:
- Biochemistry
- Immunology
- Cell Biology
Background:
- Purinergic signaling, involving extracellular nucleotides like ATP and ADP, plays a crucial role in hemostasis, inflammation, and apoptosis.
- Extracellular ATP diphosphohydrolase (ATPDase) regulates these processes by hydrolyzing extracellular ATP and ADP.
- The vascular ATPDase has been previously identified as CD39.
Purpose of the Study:
- To investigate the correlation between CD39 expression and ATPDase activity in various human cell types.
- To characterize the molecular isoforms and gene expression of CD39/ATPDase.
- To explore the potential role of coexpressed CD39/ATPDase and purinergic receptors in regulating cellular signaling.
Main Methods:
- Western blotting to detect CD39/ATPDase protein isoforms and assess expression levels.
- Northern blotting and primer extension to analyze mRNA transcripts and transcription start points.
- Detection of purinergic P2 receptor mRNAs in various human cell lines.
Main Results:
- CD39 expression levels directly correlate with ATPDase activity in human endothelial cells, platelets, monocytes, NK cells, and megakaryocytes.
- Western blotting identified one to three CD39/ATPDase isoforms, with variations attributed to post-translational modifications.
- Northern blotting and primer extension revealed two major mRNA transcripts and one transcription start point for CD39.
- Purinergic P2 receptor mRNAs were detected in all investigated cell types, indicating potential for co-regulated signaling.
Conclusions:
- CD39 is the primary vascular ATPDase, and its expression level dictates enzymatic activity.
- Post-translational modifications and distinct mRNA transcripts contribute to CD39/ATPDase heterogeneity.
- The coexpression of CD39/ATPDase and purinergic receptors suggests a regulatory role in purinergic signaling pathways.
- Modulation of CD39/ATPDase expression may offer therapeutic targets for thrombotic and inflammatory diseases.
Abstract:
Purinergic signaling may influence hemostasis, inflammatory responses and apoptosis. Therefore, hydrolysis of extracellular ATP and ADP by the ATP diphosphohydrolase (ATPDase) could regulate these processes. We have previously demonstrated the identity between the vascular ATPDase and CD39. Here we show that levels of CD39 expression correlate with ATPDase activity in human endothelial cells (EC), platelets and selected monocyte, NK, and megakaryocyte cell lines. Western blotting revealed one to three isoforms of CD39/ATPDase: mobility variations of major protein resulted from post-translational modifications. Northern blotting and primer extension indicated two major mRNA transcripts and one transcription start point, respectively. In addition, mRNAs specific for purinergic P2 receptors were detected in all of the investigated cells, suggesting that the coexpressed CD39/ATPDase may regulate purinergic signaling. Thrombotic and inflammatory responses may be modulated by the expression of CD39/ATPDase.
