Purinergic Receptor P2Y2 Control of Tissue Factor Transcription in Human Coronary Artery Endothelial Cells: New AP-1 Transcription Factor Site and Negative Regulator [Signal Transduction]

December 2nd, 2015 by Liu, Y., Zhang, L., Wang, C., Roy, S., Shen, J.

We recently reported that the P2Y2 receptor (P2Y2R) is the predominant nucleotide receptor expressed in human coronary artery endothelial cells (HCAEC), and that P2Y2R activation by ATP or UTP induces dramatic up-regulation of tissue factor (TF), a key initiator of the coagulation cascade. However, the molecular mechanism of this P2Y2R-TF axis remains unclear. Here we report the role of a newly identified AP-1 consensus sequence in TF gene promoter and its original binding components in P2Y2R regulation of TF transcription. Using bioinformatics tools, we found that a novel AP-1 site at -1363 bp of human TF promoter region is highly conserved across multiple species. Activation of P2Y2R increased TF promoter activity and mRNA expression in HCAEC. Truncation, deletion and mutation of this distal AP-1 site all significantly suppressed TF promoter activity in response to P2Y2R activation. EMSA and ChIP assays further confirmed that upon P2Y2R activation, c-Jun, ATF-2 and Fra-1, but not the typical c-Fos, bound to the new AP-1 site. In addition, loss-of-function studies using siRNAs confirmed a positive transactivation role of c-Jun and ATF-2, but unexpectedly revealed a strong negative role of Fra-1 in P2Y2R-induced TF up-regulation. Furthermore, we found that P2Y2R activation promoted ERK1/2 phosphorylation through Src, leading to Fra-1 activation while Rho/JNK mediated P2Y2R-induced activation of c-Jun and ATF-2. These findings reveal the molecular basis for P2Y GPCR control of endothelial TF expression and indicate that targeting the P2Y2R-Fra-1-TF pathway may be an attractive new strategy for controlling vascular inflammation and thrombogenicity associated with endothelial dysfunction.
  • Posted in Journal of Biological Chemistry, Publications
  • Comments Off on Purinergic Receptor P2Y2 Control of Tissue Factor Transcription in Human Coronary Artery Endothelial Cells: New AP-1 Transcription Factor Site and Negative Regulator [Signal Transduction]